CN100431707C

CN100431707C - Sample presentation device with differing wettability

Info

Publication number: CN100431707C
Application number: CNB038269201A
Authority: CN
Inventors: M·L·斯托洛维茨; C·M·贝里斯勒; D·P·帕坎; J·A·沃克二世
Original assignee: CHIYAGEN SCIENCE Co
Current assignee: CHIYAGEN SCIENCE Co
Priority date: 2003-07-14
Filing date: 2003-07-14
Publication date: 2008-11-12
Anticipated expiration: 2023-07-14
Also published as: JP2007521458A; CA2531972C; JP4668064B2; CA2531972A1; EP1656202A1; WO2005016530A1; AU2003304421A1; CN1863599A; AU2003304421B2

Abstract

The present invention relates to a sample presentation device useful in performing analytical measurement. The device can be configured to be a device that can process liquids in multiple ways, such as detention, storage, transport, concentration, positioning and diversion, and furthermore, the device can enhance the detection and the identification of analytes. The sample presentation device of the present invention is composed of one or more substrates provided with a plurality of regions with different wettability, and the present invention discloses a method for analyzing samples by using the sample presentation device of the present invention and a method for producing the sample presentation device.

Description

The different wettable sample presentation devices of tool

Background of invention

Invention field

The present invention relates to sample presentation devices useful in carrying out analysis to measure. in addition, the present invention relates to the manufacturing and the usage of this sample presentation devices.

Background

Relate to most of scientific domains requirement researchers that sample is carried out certain chemistry and bioanalysis and can identify and measure compound or the analyte (for example measurement of pesticide in the measurement of protein or the streams runoff in the blood plasma) that exists in the aqueous solution. herein, analyte is often referred to the composition of the interested fluid sample of researcher. usually, the fluid sample that will comprise analyte is by means of container (as: test tube, porous plate or cuvette) or other presentation devices (as: slide or biochip) be presented on the analysis to measure instrument. because most important purpose is to measure fast a large amount of samples (so be called sample " high flux " measurement), so with many notices be used to develop can with the standardization container and the device of automated analysis instrument logotype. for example, in the medicament research and development field, the interested researcher of screening drug candidate is often used the thousands of even millions of kinds of possible drug candidates of various analytical technologies (as: fluorescence polarization detection) screening, wherein many analytical technologies are held the sample solution that comprises drug candidate with 384 orifice plates of standard. similarly, from genomics and proteomics, drug development, clinical diagnosis is to the large-scale scientific domain of environment or biotoxin or analysis of pharmaceutical dosage forms (polluting and screening may be used for the material of bioterrorism as Evaluation Environment), and sample presentation devices has constituted very important part in researcher's the analytical equipment.

For example, in genomics and proteomics, its focus is respectively to identify and researching DNA/RNA and proteins/peptides. these fields relate to jointly to the chemistry in the organism that lives and biological partly, interaction between them and distinguish that their analytical technology carries out system research.Understand complicated life system but not the individual cells composition is the principal focal point of current biology and biomedical research in two fields. particularly, the primary and foremost purpose of genomics is order-checking and the huge database that produces whole organism gene content. compile out bacterium, yeast, the genome of nematode and fruit bat and compiled out human genome recently. similarly, proteomics is that all proteins that the specific period expresses in the pair cell is studied, its primary and foremost purpose is to obtain to be used to use the database matching instrument to identify the partially protein amino acid sequence of whole protein, this checks order different with protein fully. and the evaluation of protein permission is carried out the research (importantly identify under different condition difference expressed protein and represent the biomarker of morbid state) of protein expression and protein interaction is mapped (this helps to improve eucaryotic cell structure figure). because protein is that the main component of biological substance and the biological function of exercising nearly all key are (from conditioned reaction to transporting oxygen, again to cell and extracellular structure are provided), so it is most important that the effect of understanding protein is understood life system for us. the same with genomics, developing proteomics field has produced the information about human and other bioprotein group, though and also imperfect many these category informations of these information be stored in or be about to and be stored in the database. it is many to the life system understanding in future to estimate that we can obtain from these genomes and protein group database.

In the clinical diagnostics field, the researcher concentrates on and identifies and measure large-scale analyte. and the goal analysis thing can be actual drug candidate, in the bioavailability study of for example in clinical testing, carrying out, this research has disclosed the degree that drug candidate exists organism everywhere. alternatively, the goal analysis thing can reflect the physiological reaction that drug candidate is produced, for example in the example that whether the phosphorylation reaction product exists in measuring kinase whose enzyme reaction. because kinases is important in cell growth and breeding, so in the patient body who suffers from growth failure disease (as cancer), can be observed high-caliber kinase activity. the medicine that therefore can reduce kinase activity is possible anticancer therapeutic agent, and the existence that the analytical method that detects this type of drug candidate curative effect concentrates on the analyte of measuring kinase reaction product form usually whether. this type of and other type that important analytes in clinical diagnosis and the medicament research and development is carried out directly and indirect measurement method depend on the analytical technology that is beneficial to this measuring method and the existence of sample presentation devices.

The importance of sample presentation devices never only is confined to biomedical aspect. for example, measurement environment is polluted the interested researcher of (or educating again) degree to be needed to monitor various environmental samples, comprise water, air and pedotheque. many analytical technologies that are used to analyze this type of sample relate to the analysis to fluid sample, as sample or the pedotheque in the water quality research example, wherein said pedotheque extracts to remove multiple composition by diluting in organic and/or inorganic solvent. and therefore, can present the sample presentation devices that fluid sample analyzes is to finish the important tool of these analysis to measure.

In 9.11 worlds, back, national governments are faced with in military and domestic occasion need be easy to platform and the analytical technology that chemistry and biological reagent detect. the challenge of biological warfare investigation is comprised that sample is collected and to nontoxic and poisonous organic distinguishing. and current battlefield technology for biologic product is to utilize thermal decomposition that biologic artifact is changed into to be easier to use mass spectrometry (MS) yet the little molecule of detection., because the technology that depends on protein or peptide biomarker is more more special than at present known method, so can expect that the technology that this kind depends on protein or peptide biomarker can develop and the test of can sharing a common fate, urinalysis or blood extraction technique are united use to determine to fight the potential exposure of agent. and the purposes of independent biology sensor in battlefield and public place such as warning device makes us having great interest. and all these methods all demonstrate collects sample, preliminary treatment and carry out the challenge that sample is presented the aspect to detector.

In order to identify and measure purpose compound in the fluid sample such as DNA, RNA, protein and peptide in the serum, environmental toxin and the reagent in the environmental sample, developed multiple analytical technology. when each of these analytical technologies each has usefulness, every technology all depends on the type of the sample presentation devices that adopts at least in part. and therefore, the inherent limitation of these devices may have a negative impact for using these analytical technologies to measure the purpose compound.

In addition, manyly focus on evaluation, the analytical technology of separating or measuring analyte in the fluid sample require sample through pre-treatment step separately-promptly use particular analysis technical Analysis sample with determine that the goal analysis thing exists and quantity before handle sample. for example, numerous protein cell extraction technology produces complicated protein mixture and mixes detergent and salt, and detergent that is mixed and salt can disturb mass spectral analysis, therefore must before analysing protein, remove. current classification separation and purification process are consuming time. other purification process liquid chromatography and the gel electrophoresis of protein purification (as be used for) relates generally to volume and reclaims greater than the sample of 10 μ L, this force before analyzing, carrying out with multiple proteins detection technique (as MALDI-MS) extra concentrated. to current known analytical technology-and sample presentation devices of linking to each other with them-demand strengthened the sample purifying, sample preparation, the importance that automatic data collection and automation data are analyzed.

For example, the most frequently used and most preferred mass spectrometry is that substance assistant laser desorpted ionized mass spectrometry (MALDI-MS) .MALDI-MS is the mutation of standard laser desorb flying time mass spectrum analysis method in the proteomics field, wherein in the acidity of a large amount of molar excess, (for example absorb ultraviolet chemical matrix, nicotinic acid) there be down from the teeth outwards proteins deposited with relative HMW. this technology makes the unstable big molecule of these HMWs carry out desorb with good working condition. because mass spectrometry provides the mass accuracy of micro-example with moderate expense, responsive monitoring and rapid analysis are so it has become analysis tool important in the proteomics work.

Yet, there is number of drawbacks in MALDI-MS, especially relevant with sample preparation problem. because they can not hold the sample volume more than 2 μ L, so current MALDI-MS sample holder all is subjected to strict sample volume restriction. use volume usually up to 2 μ L, and provide diameter be 1mm-2mm do (dried-droplet) (Karas, M. and Hillenkamp, F., Anal.Chem.1988,60,2299-2301, this paper integrates as a reference). because laser only shines the sub-fraction (0.015mm that does in the one point data gatherer process ²-0.030mm ²), all be detected so can not guarantee all proteins in the sample. in addition, sample volume (up to 2 μ L) is significantly less than the volume that is reclaimed behind the conventional sample purifying, and this forces and carried out further concentrating before MALDI-MS; For example; peptide by LC and electrophoresis method purifying and protein example are recovered in the volume greater than 10 μ L usually. the result; before MALDI-MS, must carry out further concentrating to this sample. many samples also comprise detergent and the salt that disturbs mass spectral analysis, and this forces and removed these materials before MALDI-MS.

Lacking the sample homogeney is to follow another defective of MALDI-MS. when the method point sample is done in use, even make that little volume to 2 μ L also has problems because sample is heterogeneous. the sample volume of the conventional 0.5-2.0 of use μ L also carries out drying, it provides diameter is the (Karas that does of 1mm-2mm, M. and Hillenkamp, F.Anal.Chem.1988,60,2299-2301, this paper integrates as a reference). result, a little part (0.015mm in only doing ²-0.030mm ²) in the one point data gatherer process, shone by laser. unfortunately, can cause sample heterogeneity (the heterogeneity deposition of analyte) even the small size of 0.5-2.0 μ L is also known, this causes can causing peak existence, intensity, resolution ratio and mass accuracy generation marked change (Strupat, K. when laser focuses on the zones of different of doing; Karas, M.; Hillenkamp, F.Int ' l.J.Mass Spectrom.Ion Processes1991,111,89-102; Cohen, S.L. and Chait, B.T.Anal.Chem.1996,68,31-37; And Amado, F.M.L.; Domingues, P.; Santana-Marques, M.G.; Ferrer-Correia, A.J.; Tomer, K.B.Rapid Commun.Mass Spectrom.1997,11,1347-1352, this paper integrates as a reference). a large amount of single-point wave spectrums that these phenomenons cause accumulating each sample endure strict scrutiny to mass spectrometric data again. therefore, every instrument only can be analyzed a hundreds of sample every day, and usually avoids adopting automatic data acquisition.

The verified problem of sample heterogeneity that can make when spot diameter is reduced to the order of magnitude of laser diameter minimizes. and in this case, most of sample can be shone simultaneously, has therefore improved sensitivity and repeatability (Little, D.P.; Cornish, T.J.; ODonnell, M.J.; Braun, A.; Cotter, R.J.; Koster, H.Proc.Natl.Acad.Sci.U.S.A.1997,69,4540-4546; And Gobom, J.; Nordhoff, E.; Mirgorodskaya, E.; Ekman, R.; Roepstorff, P.J.Mass Spectrom.1999,34,105-116, this paper integrates as a reference). at U.S. Patent number 6,287, the sample holder of describing in 872 is further described (Schuerenberg, M.; Lubbert, C.; Eickhoff, H.; Kalkum, M.; Lehrach, H; Nordhoff, E.Anal.Chem.2000,72,3436-3442, this paper integrates as a reference), wherein showing analyte restriction is deposited into and not only reduced the problem that accompanies with the sample heterogeneity in the speckle diameter, and obviously improved detection sensitivity. its defective is the spot size that will obtain this expection, sample volume must be reduced to be lower than 2 μ L.

In order to overcome these sample volumes and impurity problem, the researcher has used the sample holder through designing or has been used for the small-sized post of sample pretreatment. and the example of this type of sample holder is that commerce can get, as the AnchorChip from Bruker Daltonics GmbH ^TM. this AnchorChip ^TMThe sensitivity of product by improving MALDI-MS at the position concentrating sample that accurately limits, and particularly, this product comprises the non-wettability hydrophobic material thin layer that carries the hydrophilic lattice array of wettable. with AnchorChip ^TMUsing main limitation together is that the liquid sample volume that requirement is applied to each anchor point (anchor) is limited in 0.5 μ L-3.0 μ L (about AnchorChip ^TMArticle one in 11 general rules of the sample preparation of target is seen AnchorChip ^TMTechnology, the 1.6th edition, Bruker DaltonicsGmbH, in November, 2000, this paper integrates as a reference); The volume restrictions that the example that this manufacturer provides in the document of this product further drips fluid sample or 0.5 μ L or 1.0 another limitation of μ L. be that analyte and pollutant (salt, detergent) are concentrated through the laser irradiation area territory of being everlasting.Therefore, as mentioned above, sample at first must desalination before being applied on the mass spectrograph sample holder and/or

Or concentrate (production of Millipor company on the similar small-sized post sample preparation device

Be the micro-column that is used for sample concentration and desalination, it prepares (Rusconi, F. by pipetting point with reverse chromatography media filling pipette; Schmitter, J.-M.; Rossier, J.; Le Maire, M.Anal.Chem.1998,70,3046-3052, this paper integrates as a reference)) yet., use homemade micro-column or commercial

Be consuming time, cost is quite high, and proof be difficult to automation and can only realize the medium recovery of specimen material usually. therefore, AnchorChip ^TMHave and the relevant many identical limitation of current other MALDI-MS sample holder.

Developed the MALDI-MS alternative technique that is used for blood serum sample protein analysis (protein profiling). this technology is called surface-enhanced laser desorption ionization mass spectrometry (SELDI-MS), and it is finding to obtain the result aspect differentiation of oophoroma biomarker and prostate cancer and the benign prostatic hyperplasis biomarker. when carrying out SELDI-MS, at first analyte optionally is trapped on the sample holder as the affinity capture device, wherein said sample holder has functionalized surface. then catch the site by laser desorption with the analyte ionization of being detained so that can detect to them, it does not need to require as other LC-mass spectrometry combination method to realize that they are described in U.S. Patent number 5 from the recovery .SELDI-MS that is detained the surface, 719,060,5,894,063,6,020,208,6,027,942,6,124,137,6,225,047 and 6,579,719, this paper all integrate as with reference to no matter recently results reported how, the SELDI-MS method goes wrong in the operating process of being everlasting, and this is because be not best on the surface of being detained the best aspect the biological analyte aspect the presenting of analyte in the laser desorption ionisation process.

Also used other to be used to separate technology with the analyte of purifying such as protein. for example, undertaken by the technology consuming time of two dimensional gel electrophore-sis and multidimensional LC that the biological sample classification separates and the method for purifying is well-known, it is faster, the technology of muting sensitivity, as the expendable post or the pipette that have the chromatography bed pipette point. the gel electrophoresis that is used for the isolated protein mixture can be one dimension or two-dimentional. one dimension gel electrophoresis (being also referred to as SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis)), protein mixture only separates by their molecular weight. in two-dimensional gel electrophoresis (being also referred to as 2D-PAGE), mixture separates by their isoelectric point earlier, molecular weight by them separates subsequently. and the resolution ratio that a defective of this technology is the method is low, be that each is differentiated point and may comprise more than one protein. another shortcoming is that the dyestuff that is used to observe separation can not dye to all proteins. LC (LC) with " high performance liquid chroma-tography " (HPLC) or " multidimensional LC " (if having used a more than chromatographic column) be usually to utilize different post chemical actions as the advantage of .LC that people are known. compare with the gel electrophoresis that can not effectively separate littler peptide, LC can be used for from enzymic digestion thing isolated peptides mixture. and solid phase extraction (SPE) provides the purifying of shortcut and its to be used for many fields of gathering from the organic synthesis to the environmental sample. and this method is faster than liquid-liquid extraction method or HPLC, it consumes the solvent of less amount and can be used for providing multiple device from gas or fluid sample extraction of analytes .SPE technology, enumerates some and pipettes point as pipette, post, film and 384-orifice plate.

In the medicament research and development process, also develop other sample presentation devices of in known analysis methods, using. for example, ADMET (absorbs, distribute, metabolism, drain, toxicity) the Empore card (http://www.3m.com/empore) that uses in the research, this is C18 RP (oppositely) adsorbent that is embedded in the film, it claims the number of times that can reduce the sample purification step, and because the sample that loads keeps dry, so can also file and concentrate. the sample purifying needs three steps: load sample is to card, if card is transferred to elution instrument and 100% sample directly is eluted in the mass spectrograph. elution volume can be low as far as possible, then the Empore card can be used for peptide digest sample is loaded into MS, otherwise the low concentration peptide will be lower than detection limit.

Therefore, need a kind of like this sample presentation devices, it can connect various analytical methods so that high-sensitivity detection biological and chemical part. in addition, also need a kind of like this sample presentation devices, itself and the conventional sample volume compatibility that reclaims from LC and electrophoretic separation and other separation/purification technology, the fluid sample that will comprise analyte points to institute's restricted area so that the problem that accompanies with the sample heterogeneity is minimized, if cause detection sensitivity to increase. can obtain this kind sample presentation devices, then can realize the automation of sample treatment, for example, the standardization porous plate processor and the liquid handling robot of life science industry. more importantly, they can also directly be collected the chromatography eluate and analyze with MALDI-MS subsequently. and these abilities will improve jointly uses multiple analytical technology well known by persons skilled in the art that biological and chemical is partly carried out flux detection and measurement. and these benefits of the present invention and other benefit are described in more detail hereinafter.

Summary of the invention

For for the known sample presentation devices of using in the various analytical methods of identifying chemical individual and organism, sample presentation devices of the present invention provides attractive alternative. and in addition, the invention provides the method for making this sample presentation devices and use them that the analyte that is contained in the fluid sample is carried out the method for analysis to measure on a large scale. the character of sample presentation devices uniqueness of the present invention has been to overcome many shortcomings (as mentioned above) of following known analytical technology and sample presentation devices that links to each other with them or container.

Such as genomics, proteomics, medicament research and development, clinical diagnostics, biology sensor and environmental toxin and reagent detection range, be used to identify that the technology of chemistry and biological part is a mass spectrometry, wherein can only obtain minute quantity sample and expectation usually carries out quick flux to a large amount of samples and handles. other analyte detection method such as fluorescence polarization, immunofluorescence spectroscopic methodology, gel chromatography, ion exchange chromatography, affinity chromatography also can detect as the high flux of biological and chemical part, therefore also can with sample presentation devices coupling of the present invention.

Sample presentation devices of the present invention provides the attractive alternative of the known sample presentation devices of using in various analysis. and for example, the present invention makes volume optionally be detained up to the analyte of 100 μ L and concentrates on the biochip surface.In addition, since analyte be from be designed to basically not in conjunction with or the part of the sample presentation devices of opposing combination detect, so compare with directly detecting on based on the affinity capture apparatus surface of biochip or on its surface analyte is had directly to detect on other sample presentation devices surface of obvious compatibility, its detection sensitivity is higher.

Because the sample presentation devices of invention only needs once fluid operated in preferred embodiments, so further making, the present invention follow analyte to minimize from the potential loss that a surface is transferred to another surface process. this character with sample presentation devices surface opposing analyte causes the reduction of goal analysis thing loss, and in known method, there is the loss of this goal analysis thing. different with SELDI-MS, the present invention does not relate to the desorption process of catching site of institute's bound analyte from the affinity capture device, and be to use a kind of like this sample presentation devices, but wherein analyte is it not to be had the perception compatibility or do not have the surface of combination to carry out desorb from analyte.

In addition, fluid sample can be with controlled manner at the enterprising line operate of sample presentation devices of the present invention and mobile. and this makes sample can concentrate in analyte and the uncombined basically analyzed area of sample presentation devices. and, this sample that also allows to comprise analyte moves to surperficial zones of different, wherein each zone has different character for analyte, this makes and can carry out purifying to analyte before detecting, separate and/or modification. in addition, the present invention relates to such sample presentation devices, the character of its surperficial different piece can respond different chemical or physical stimulation (as, heat, ultra-violet radiation) changes, so that can in the sample treatment process, handle this surface nature for analyte.This kind variation of surface nature can be designed to reversible or irreversible.

These character and other feature of sample presentation devices of the present invention are described in more detail hereinafter. and the present invention comprises the method for sample presentation devices, perparation of specimen presentation devices and the method for using sample presentation devices.

Sample presentation devices

The present invention relates to sample presentation devices useful in carrying out analysis to measure. in one embodiment, the present invention relates to have one or more for various samples to be analyzed the sample presentation devices of wettable zones of different. these different wettables zones cause between the zone be detained, concentrate different with the ability of analyte in the moving liquid sample. these zones can be multiple shape and size, and can be continuous each other or discontinuous.

Sample presentation devices of the present invention can be made up of zones of different, one of them zone is being best aspect the delay fluid sample. sample presentation devices of the present invention can also comprise different wettable zones, and one of them zone is being best aspect the high-sensitivity detection of analyte.

Sample presentation devices of the present invention can comprise two dimension or three-dimensional surface, and wherein each surface all has two or more different wettable zones.

Sample presentation devices of the present invention comprises the substrate that can make of multiple material, these materials including, but not limited to for example glass, semiconductor, metal, polymer (for example plastics) and other hydroxylating material as having SiO ₂The layer silicon, have Al ₂O ₃The aluminium of layer etc. substrate preferable alloy (for example gold) or semiconductor (for example silicon).

Sample presentation devices of the present invention also comprises such substrate, promptly it has been carried out finishing by the known method of those of ordinary skills in order to produce multiple zone at substrate surface, these zones have heterogeneity aspect wettable. and this kind finishing is including, but not limited to basad interpolation self-assembled monolayer (SAM), polymer (linear or branch) and Langmuir-Blodgett assemble thing. and with SAM is example, after it is added to substrate, SAM has just formed the sample presentation devices surface that fluid sample will contact. depend on the composition of the special SAM that uses, the surface of sample presentation devices of the present invention has different character aspect the analyte compatibility (or shortage compatibility) aspect the wettable and in to fluid sample. and can SAM be added on the sample presentation devices of the present invention in the mode that produces zones of different, wherein its character has reflected that employed SAM. other surface modification technology well known by persons skilled in the art also is contained among the present invention in the specific region.

The regional kind that can comprise as for this sample presentation devices surface, they mainly are to rely on them that the different wettables of analyzed sample are characterized, conversely, wettable difference has caused again producing is detained or in conjunction with the different zone of analyte ability in the fluid sample. these zones broadly are called " frontier district ", " liquid holding zone " and " analysis area " though. the present invention also considers to comprise the zone of two or more types, but the present invention only needs to exist two types zone. the present invention can also comprise more than one zone in each kind-for example, sample presentation devices can comprise a plurality of liquid holding zones, and wherein each zone is for fluid sample and/or wherein comprise the analyte different in kind.

First kind zone is called as " frontier district ", and it comprises and treats the not wettable basically zone of analytic sample. and compare with other zone, the frontier district is the zone with the highest sample contact angle.

For sample to be analyzed, higher relatively (and compare wettable with analysis area relative relatively poor than frontier district wettable for the second class zone (being called as " liquid holding zone ") wettable, following). the contact angle of liquid holding zone is lower than the contact angle of frontier district relatively, and (and contact angle is relatively higher than the contact angle of analysis area, following). originally liquid holding zone can also have the contact angle that is equal to or less than analysis area, but because chemistry or physical stimulation make liquid holding zone present the contact angle higher than the analysis area before chemistry or the physical stimulation, so just cause fluid sample has been pointed to than another preferred zone, zone.

Liquid holdup can be divided into two subclass. in a subclass, liquid holding zone is designed to be able to realize the purpose of fluid sample delay, the combination of resisting analyte again basically simultaneously. in second subclass, liquid holding zone is designed to not only can be detained fluid sample but also can be basically in conjunction with the analyte in the fluid sample, because can catching analyte, it therefore it is called " trapping region ". this second subclass can also comprise a kind of like this surface, promptly can sizable bound analyte, but when being subjected to such as ultra-violet radiation, become uncombined basically surface behind the chemistry of electric current or heat or the physical stimulation.

The 3rd class zone is called as " analysis area ", and it is the zone of comparing the wettable maximum (and having minimum contact angle) for sample with other zone. and analysis area is configured to analyte in conjunction with opposing. can optimize analysis area so that improve sensitivity for analysis according to size, shape and surface nature to the goal analysis thing.

The liquid capacity of sample presentation devices of the present invention depends on the size in zone. for the border circular areas of 3mm diameter, its liquid capacity can hold the fluid sample of this quantity and need not physical boundary, reservoir or hole up to about 100 these sample presentation devices of μ L.. various zones can be accurately positioned, be convenient on multiple analytical instrument such as the mass spectrometer the high flux automatics or with its compatibility.

In another embodiment of sample presentation devices of the present invention, sample presentation devices is called as " target chip (target chip) ", be abbreviated as Tn, wherein " n " is the figure notation about the quantity of zones of different on the sample presentation devices surface, and wherein " n " can be to the Any Digit between the infinity from 2. therefore, for example, T2 target chip has two zones, T3 target chip has three zones, or the like. the present invention has considered to comprise many sample presentation devices more than 2 or 3 zones, and the present invention never in any form with region limits in specific quantity. along with the increase of region quantity, total effect approaches gradient. the target chip is by the sample presentation devices that is designed to one or more zones of opposing analyte combination are formed.

For example, as for T2 target chip, this sample presentation devices comprises two zones-be frontier district and analysis area. with the region surface of contact liq sample be designed to analyze thing in conjunction with opposing-be that analysis area is that analyte is in conjunction with opposing. during the drying steps before analyzing, the region surface of contact liq sample has limited analyte effectively.

As for T3 target chip, sample presentation devices comprises three zones-be frontier district, liquid holding zone and analysis area. with the region surface of contact liq sample be designed to analyze thing in conjunction with opposing-be that liquid holding zone and analysis area are that analyte is in conjunction with opposing. during drying steps, the region surface of contact liq sample is concentrated into analysis area with analyte effectively.

Therefore, sample presentation devices of the present invention can comprise different zones, and wherein each region list reveals the MIN absorption of analyte.

In another embodiment of sample presentation devices of the present invention, sample presentation devices can be called " catching chip " or " catching/concentrate chip ", and be abbreviated as Xn, wherein " n " is the figure notation about region quantity on the sample presentation devices surface, and " n " can be to the Any Digit between the infinity from 2. therefore, for example, X2 catches chip and has two zones, X3 catches chip and has three zones, or the like. the present invention has considered to comprise many sample presentation devices more than 2 or 3 zones, and the present invention is confined to specific quantity with the zone never in any form. along with the increase of region quantity, total effect approaches gradient. and catch chip and catch/concentrate the sample presentation devices that chip is made up of the one or more zones that are designed to bound analyte.

For example, catch chip as for X2, sample presentation devices comprises two zones-be frontier district and trapping region. based on the chemistry on trapping region surface or biological property the region surface of contact liq sample is designed to be able to catch analyte-be the trapping region bound analyte. and during the drying steps before analyzing, the region surface of contact liq sample has limited analyte effectively.

Catch/concentrate chip as for X3, sample presentation devices comprises three zones-be the frontier district, trapping region and analysis area. the frontier district is designed to not wettable basically. trapping region is designed to be able to catch and bound analyte. analysis area is designed to analyze thing in conjunction with opposing. before analyzing by one of multiple known analyzing detecting method, analyte is being shifted between trapping region and the analysis area. during the drying steps before analyzing, analyte has been limited on the analysis area surface that comprises fluid sample effectively. and utilize the character on trapping region surface can realize the transfer of fluid sample from the trapping region to the analysis area-promptly, if trapping region is lower than the wettable degree of analysis area, do not need the physical interventions fluid sample will move to analysis area so from trapping region yet. alternatively, also can design trapping region, make its character can respond chemistry or physical stimulation (as heat, ultra-violet radiation) changes, thereby cause the wettable degree of trapping region to be lower than analysis area, and therefore cause fluid sample to move to analysis area from trapping region.

In another embodiment of sample presentation devices of the present invention, sample presentation devices can be above-mentioned target chip and the combination of catching chip. in this embodiment, sample presentation devices is by the surface composition with different functionalities. and the sample presentation devices of these types relates to by mechanical means (as pipetting by pipette) or other method (as by wettable difference between the zone) fluid sample from a zone-transfer to another zone. as an example, " catch-shift-concentrate chip " and (be abbreviated as the sample presentation devices that X2-shift-T3) is made up of X2 chip and T3 chip, wherein said X2 chip is made up of two zones (being frontier district and trapping region), and described T3 chip (is the frontier district by three zones, liquid holding zone and analysis area) to form. the transfer of analyte (mechanical transfer or other) betides between the liquid holding zone of the trapping region of X2 chip and T3 chip. in addition, relating to trapping region can also be used for before the analyte to fluid sample detects it being separated with the embodiment of the sample presentation devices of liquid holding zone associating with associated form, concentrate, purifying and modification. therefore, for example, fluid sample can be placed on the T2 chip so that the analyte of sample is limited in analysis area.Then, those samples are transferred to comprise the frontier district, the X3 chip of trapping region and analysis area. in this example, trapping region can be designed to be able in conjunction with the lipid part in (and therefore removing) fluid sample, so that when with sample application during in the X3 chip, it moves to trapping region (it has the wettable of higher degree) from the frontier district, lipid in the sample partly is incorporated into the trapping region surface, and remaining sample moves to analysis area (because it has the wettable of top). in this example, fluid sample is limited on the T2 chip, and lipid moves on the X3 chip subsequently, so that the final sample of analysis area analysis be concentrate and be that purifying becomes no lipid. owing to trapping region being designed in conjunction with multiple different analytes, and can use any multiple combination in these zones, have purifying on a large scale so can create, concentrate, the sample presentation devices of separation and modification ability (about one or more analytes).

Fluid sample can be changed from the mechanism that a sample presentation devices is transferred to another sample presentation devices. use above example, mechanically (as pipetting) by pipette remove the sample that concentrates from T2, and place on the X3 sample presentation devices separately. alternatively, connect T2 and X3 sample presentation devices by a zone, this regional wettable can respond chemistry or physical stimulation (as ultra-violet radiation) and change, so that when the zone between T2 sample presentation devices analysis area and the X3 device trapping region is exposed to ultra-violet radiation, will make this regional wettable be higher than T2 device analysis district but be lower than X3 device trapping region, therefore the sample that concentrates in T2 sample presentation devices analysis area just is transferred to the trapping region of X3 device, thereby sample is transferred to X3. again from T2, and the surface of utilization enormous quantity (the different wettable of tool with different analyte in conjunction with character) and configuration thereof can be created has purifying on a large scale, concentrate, the sample presentation devices of separation and modification ability (about one or more analytes).

Sample presentation devices of the present invention also provides the zone of the different wettables with difformity or pattern. for example, in one embodiment, sample presentation devices has the zone of concentric circles form, middle section is an analysis area, analysis area is surrounded by liquid holding zone, and liquid holding zone is surrounded by the frontier district. owing to can use multiple light pattern technology to produce the zone, and because known light pattern technology can provide the very big variation of gained pattern, so large-scale possibility shape can be arranged for multiple zone, pattern and configuration. in addition, perhaps the multiple character in different wettables zone can produce can point to analyte the surface and go up single or multiple appointments or pre-determine position (addressable sites for example, road or zone) sample presentation devices.

Sample presentation devices of the present invention is suitable for the processing of biological and abiotic fluid sample. and they also are fit to be applied in the analyte detection method of wide scope, and these methods for example (include but not limited to) other known analysis methods of analyte in mass spectrometry, multiple chromatography method, immunofluorescence spectroscopic methodology and detection and the measurement fluid sample.

Each of above-mentioned variation all is designed to allow maximum flexibility ratio in the design and use of sample presentation devices, described sample presentation devices have than known method stronger present the ability that analyte detects and analyzes. therefore, sample presentation devices of the present invention has guides ability to the analysis area that is designed to be able to improve the analyte high-sensitivity detection with analyte. and therefore, sample presentation devices of the present invention provides the analyte that improves deposition.

The manufacturing of sample presentation devices

The present invention also has other embodiment to comprise the method for producing or making above-mentioned sample presentation devices.

Form by self-assembled monolayer (SAM) in the embodiment of (wherein depend on the difference self-assembled monolayer between the SAM that uses and formed different zones) on the surface, sample presentation devices of the present invention comprises the multiple SAM zone that produces by known light pattern technology. and therefore, the present invention comprises that also use makes the method for the sample presentation devices of being made up of SAM as a kind of light pattern technology of method for optimizing.

The substrate surface of sample presentation devices of the present invention is generally modified or patterning by method known to those skilled in the art. as an example, the surface of substrate can be modified or patterning by using self-assembled monolayer (SAM), wherein the surface of sample presentation devices substrate is modified and the surface of its exposure gives substrate special chemical property with self-assembled monolayer. for special substrate is selected to comprise 1 °, 2 °, the multiple SAM of 3 ° or 4 ° compositions, this provides unique surface characteristics and character for substrate surface. particularly, use multiple SAM and make base patternization, so that it comprises a plurality of zones that each zone has different surfaces characteristic and character. with the method for SAM patterning is known in the art, and comprises the ultraviolet patterning, the photolithography patterning, little seal (microstamping), electron beam patterning and reactive ion etching method.

The zone that produces on substrate surface can be an arbitrary shape, circular. in addition, the zone regional with other or continuous or discontinuous-be the zone can all adjacent one another are or one or more zones and one or more other zones non-conterminous. the zone that produces on the sample presentation devices substrate surface preferably has treats the different a plurality of zones of analytic sample wettable.

As another embodiment of the present invention, it provides the method for the accurate positioning analysis thing of energy with the sample presentation devices of convenient automatic data collection of making.

The purposes of sample presentation devices and application

In another embodiment, sample presentation devices of the present invention with multiple analytical technology and method coupling in have a lot of purposes. therefore, the present invention includes the method for using above-mentioned sample presentation devices. more specifically, the present invention includes and use sample presentation devices of the present invention or on the sample presentation devices or on a plurality of sample presentation devices, identify the existence of analyte in the sample and the method for analyzing a plurality of samples.

In fact, can detect, identify or measure that any method of analyte may be used to and sample presentation devices coupling of the present invention in the fluid sample. the example of this type of analytical method includes but not limited to that MALDI-MS or this sample presentation devices of electron spray ionisation MS. especially are fit to unite use with the high throughput analysis measuring technique, for example in MALDI-MS, use, wherein the sample presentation devices analysis area be configured in the mode that promotes the high flux data acquisition.

Sample presentation devices of the present invention can also be used for the operating liquid sample and wherein comprise analyte. the different wettable characteristics and the capture characteristic that can be designed to have based on the sample presentation devices surface, the sample presentation devices design can be used for operating, concentrate, the location, store, shift (by with do not pass through mechanical intervention), reclaim (by or do not pass through mechanical intervention), analyze, modify or processing (modifying reagent) by operational analysis thing on sample presentation devices, perhaps classification separating liquid sample or wherein comprise analyte. in addition, owing to sample presentation devices of the present invention can be designed to be able to respond chemistry or physical stimulation (as heat, ultra-violet radiation, pressure, electromagnetic radiation) finishes any function of these functions, so sample presentation devices of the present invention can reversible or irreversibly realize these functions, and can response external pressure further finish the multiple combination of these functions.

Any fluid sample (with analyte) can be connected use with sample presentation devices of the present invention. for example, the present invention can be used for analyzing the fraction that reclaims from LC. and the present invention can be used in analysis from the protein spots of 2D gel electrophoresis cutting-out or the enzymic digestion thing for preparing from the fraction that affinity chromatography (being ICAT (isotope affinity tag)) is collected. and the present invention can also be used to analyze the sample that reclaims from biology sensor. and the present invention can also be used to use the porous form Robotics of standard and analytic approach to carry out sample transfer in 1: 1. really, sample presentation devices of the present invention can be used for handling and operating the fluid sample that may originate and obtain from any, and no matter this sample is laboratory experiment result (for example enzymic digestion thing and the biology sensor sample example of above identifying), still the water sample of river (for example from) that from environment, obtains, still direct (for example human urine sample) that obtains from live organism.

The present invention can also be used to the file purpose or store for further analyzing the sample that carries out. in other words, need after being transferred to analysis area, fluid sample not carry out immediately the detection and the analysis that comprise analyte in the fluid sample.

Therefore, multiple embodiments of the present invention provides the sample presentation devices that can play multiple liquid handling function, and wherein these functions include but not limited to sample/analyte processing and liquid deposition, delay, transfer, locate and relocate and store.

Feature and advantage

The present invention also comprises following extra feature and advantage at least except described many features of top summary of the invention part and advantage:

Detection is present in the analytical method (as MALDI-MS) of analyte in the fluid sample can not carried out on the single surface of bound analyte basically, and this causes, and sensitivity for analysis increases, reproducibility of results increases and be comparable from the result of different trapping regions.

Handle about sample liquids, it can analyze the sample volume (can reach about 100 μ L for the 3mm diameter region) that has increased, picture on surface can be turned to the have SBS gauge orifice format (i.e. 96/384/1536 hole format) of (biomolecular screening association (Society for Biomolecular Screening)), and therefore it can join with machine in normal service people technology and other method for high-flux analysis.

It can make the flux of various analysis (for example MALDI-MS) increase, because accurately placed in order to carry out high-throughout data acquisition zone. and as for MALDI-MS, its analysis area is that best size is (promptly less than 2mm ², and preferably less than 1mm ²). its sample/matrix has been improved crystallization, and this causes having improved analysis area internal ionization uniformity. and compare with doing analysis, it has littler analysis area, causes detected area littler, and this can realize high throughput analysis.

Sample presentation devices of the present invention can be by concentrating analyte in the sample of analyzing dilution at analysis area.

Can not need multiple separating step and analyte in the separating liquid sample, multiple separating step is such as analyte being incorporated into the separate analytes from the post of having to then on the ion exchange column in elution step subsequently. and really, the SAM (being designed in conjunction with different analytes) that has the different surfaces chemical property by use can separate and the purifying specific analyte with high specificity.

Can handle large quantities of fluid samples and analyte by sample presentation devices of the present invention; this has been avoided the defective of above-mentioned known presentation devices and analytical method. and as described below; though sample presentation devices of the present invention is particularly useful for proteomics field and laser desorption ionisation mass spectrometry, the purposes of the device of asking for protection only is confined to that field never in any form.

The accompanying drawing summary

Above-mentioned target of the present invention and other target are become obviously, wherein:

Fig. 1 a has described sample presentation devices of the present invention, wherein Zhong Yang analysis area and liquid holding zone on every side be for concentric each other, and wherein liquid holding zone is surrounded by the frontier district. and what Fig. 1 b described is the cross-sectional view strength of the sample presentation devices of describing among Fig. 1 a.

Fig. 2 has described the surface of sample presentation devices of the present invention, wherein should further form on the surface by 16 pairs of analysis areas and liquid holding zone, wherein said analysis area and liquid holding zone are for concentric each other, and wherein paired analysis area and liquid holding zone are surrounded by common frontier district. in this case, this sample presentation devices is according to organizing corresponding to the geometry of standard 96 orifice plates.

Fig. 3 has described the surface of sample presentation devices of the present invention, wherein partial analysis district and partially liq stagnant area are adjacent one another are, wherein the part not adjacent to each other of analysis area and liquid holding zone is surrounded by common frontier district, and wherein the surface area of analysis area less than the surface area of liquid holding zone.

Fig. 4 a has described the surface of sample presentation devices of the present invention, wherein the analysis area shaped design is become to be easy to the automation collection of mass spectrometric data. and Fig. 4 b has described the enlarged drawing of analysis area, is presented to be measured as about 100 μ m ²In 36 zones, and these zones corresponding the single zone of in mass spectrometry, being taken a sample by laser.

Fig. 5 has described the surface of sample presentation devices of the present invention, wherein should further form on the surface by 96 pairs of analysis areas and liquid holding zone, wherein analysis area and liquid holding zone are for concentric each other, and wherein paired analysis area and liquid holding zone are surrounded by common frontier district. in this case, this sample presentation devices is according to organizing corresponding to the geometry of standard 96-orifice plate. liquid holding zone is elongated so that the liquid containing ability is maximum and make the distance between the adjacent domain minimum. and the sinuous pattern that covers on preceding two rows of sample presentation devices has shown the route of being described by the liquid stream deposition of chromatography eluent in automation fraction collection process.

Fig. 6 a to Fig. 6 h has illustrated and has made the related step of sample presentation devices of the present invention, and wherein the alkyl hydrosulfide on the gold (alkylthiol) is used for finishing and the ultraviolet patterning techniques is used for patterned surface.

Fig. 7 a to Fig. 7 l has illustrated and has made the related step of sample presentation devices of the present invention, and wherein the alkyl hydrosulfide on the gold is used for finishing and photolithography is used for patterned surface.

Fig. 8 a to Fig. 8 l has illustrated and has made the related step of sample presentation devices of the present invention, and wherein the alkyl silane on the silicon (alkylsilanes) is used for finishing and photolithography is used for patterned surface.

Fig. 9 a to Fig. 9 f has described the different phase in the processing procedure, and the water-soluble sample of large volume that is deposited on sample presentation devices of the present invention surface thus is at the regional inner drying corresponding to analysis area.

Figure 10 a to Figure 10 d has described not have with the tool liquid holding zone the relevant surface and the droplet drying characteristic of sample presentation devices of analysis area. and Figure 10 e to Figure 10 h has described the relevant surface and the droplet drying characteristic of sample presentation devices of the no liquid stagnant area with the tool analysis area.

Figure 11 a to Figure 11 h has described the image that video contact angle device is write down during the lip-deep droplet drying of sample presentation devices of the present invention, is that 0.6mm and liquid holding zone diameter are 1.5mm. through Measurement and analysis district diameter wherein

Figure 12 is the curve map of summing up contact angle, drop width and the drop height relevant with institute's rendering image among Figure 11 a to Figure 11 h.

Figure 13 is the photo of sample presentation devices of the present invention that deposits the liquid of 5 μ L-70 μ L volumes thereon.

Figure 14 a is the photo that 5 μ L-40 μ L liquid depositions are gathered after on the sample presentation devices of the present invention immediately. and each drop comprises the alpha-cyano-4-hydroxycinnamic acid (HCCA) of equal amount. and Figure 14 b is that drying is concentrated and guides the photo to the HCCA of analysis area on the sample presentation devices of being described in Figure 14 a owing to sample. seeing reference of concentricity district made an addition to the top of dry HCCA.

To detailed description of the preferred embodiments

Definition

Unless otherwise defined, all technology used herein and scientific terminology have the meaning of this field that the present invention belongs to technical staff common sense. and as used herein, following term has their implication of ownership, unless otherwise noted.

" analyte " refers to the component of desired test sample. this term can refer to one-component or the various ingredients in the sample.

" sample " refers to be presented on the lip-deep any material that comes from biological or abiotic source of sample presentation devices. this sample can be original with it, the form that is untreated and/or handle after be applied to sample presentation devices, wherein said processing includes but not limited to modifications, classification separation, extracting and concentrated. sample of the present invention can be liquid or on-liquid sample.

" substrate " refers to present or the material of support surface.

" surface " refers to the outside or the upper bound of main body or substrate.

" not combination basically " or " in conjunction with opposing " or " analyte is in conjunction with opposing " refer to some surperficial characteristic relevant with sample presentation devices of the present invention, though but it does not show the perception compatibility or does not show analyte and surperficial combining. some combinations may take place, and can carry out particular design so that will be in conjunction with being reduced to the level that is lower than used analytical method detectability to these surfaces.

" surface tension " refers to the character of liquid, wherein is deposited on lip-deep drop owing to the force unbalance of near surface molecule inner injection trends towards being shrunk to minimum possible contact area.

" wettable " refers to that the surface of solids is by the wetting degree of fluid sample. unless otherwise noted, fluid sample is water miscible in nature.

" contact angle " refer to surface of solids plane and originate from three contact (solid/liquid/steam) point drop border tangent line between angle.

" matrix " refers to the material that uses in the analytical technique of mass spectrum such as MALDI-MS or SELDI-MS, it is used to absorb the energy of laser and shifts this energy to analyte molecule, molecule can ionization greatly to make instability. in SELDI-MS, this matrix is called " EAM " or " energy absorption molecule ". and often the reagent that uses as the matrix of biological analyte detection includes but not limited to trans-3,5-dimethoxy-4 '-hydroxycinnamic acid (sinapic acid, SA), alpha-cyano-4-hydroxycinnamic acid (HCCA) and 2,5-dihydroxy-benzoic acid (DHBA). other suitable matrix is known to those skilled in the art.

" SAM " refers to that self-assembled monolayer .SAM is by suitable substrate being immersed in the solution that is dissolved in the active surfactant in the organic solvent and the molecule assemblage of spontaneous formation.

Description to sample presentation devices of the present invention

Provide than the above more detailed introduction of invention summary in following description sample presentation devices of the present invention. and, sample presentation devices of the present invention by relating to accompanying drawing, sample presentation devices of the present invention manufacture method and the purposes and the application (wherein each is describing in detail down) of sample presentation devices of the present invention further describe.

Mention as top institute, for for the known sample presentation devices of using in the various analytical methods of chemical individual and organism evaluation, sample presentation devices of the present invention provides attractive alternative. and in addition, the invention provides the method for making this sample presentation devices and use them that the analyte that is contained in the fluid sample is carried out the method for analysis to measure on a large scale. the peculiar property of sample presentation devices of the present invention has overcome follows known analytical technology and the sample presentation devices of use or the many shortcomings (being described in the foregoing background part) of container of linking to each other with them.

More specifically, sample presentation devices of the present invention provides the attractive alternative of the known sample presentation devices of using in wide surface analysis method. and they have extra benefit, for example, it makes analyte in the fluid sample be detained with the volume selectivity of 100 μ L nearly and concentrates in biochip surface. in addition, since analyte be from be designed to basically not in conjunction with or the part of the sample presentation devices of opposing combination detect, so with on based on the affinity capture apparatus surface of biochip or the lip-deep direct detection that analyte had other sample presentation devices of obvious compatibility of its surface compare, its detection sensitivity is higher.

Because sample presentation devices of the present invention only needs once fluid operatedly in preferred embodiments, follows analyte to minimize from the potential loss that a surface is transferred to another surface process so the present invention further makes.

In addition, because analyte is as for example being not joined on the affinity capture device in the SELDI-MS biochip, so fluid sample can be with controlled manner at the sample presentation devices enterprising line operate in surface of the present invention and mobile. this makes sample can concentrate in the analyte analysis area of uncombined sample presentation devices basically. and, this sample that also allows to comprise analyte moves to lip-deep zones of different, wherein each zone has different characteristics for analyte, and this makes and can carry out purifying, separation and/or modification to analyte before detecting.

The present invention relates to such sample presentation devices, be that the character of its surperficial different piece (for example can respond number of chemical or physical stimulation, heat, ultra-violet radiation) and change, so that can in the sample treatment process, handle this kind surface nature for analyte. this kind variation of surface nature can be designed to reversible or irreversible.

Therefore, the present invention relates to be used to carry out the sample presentation devices of analysis to measure. in one embodiment, the present invention relates to such sample presentation devices, it is one or more for the different different zones of sample wettable to be analyzed to be that its surface has. and have different wettable these zones and cause having produced delay, concentrate with the moving liquid sample in the different zone of analyte ability. these zones can be multiple shape and size, and can be continuous mutually or discontinuous. sample presentation devices of the present invention can comprise two dimension or three-dimensional surface, and wherein each surface all has the wettable zone of two or more differences.

Sample presentation devices of the present invention comprises the substrate that can make of multiple material, and these materials for example include but not limited to, glass, silicate, semiconductor, metal, polymer (for example plastics) and other is such as the SiO on the silicon ₂, the Al on the aluminium ₂O ₃Deng the hydroxylating material. preferably, substrate is metal (for example gold) or semiconductor (for example silicon). sample presentation devices of the present invention also comprises so a kind of substrate, promptly carried out finishing by the known method of those of ordinary skills in order to produce multiple zone at substrate surface, these zones have heterogeneity aspect the wettable. and this kind finishing includes but not limited to add self-assembled monolayer (SAM) in substrate, polymer (linear or branch) and Langmuir-Blodgett assemble thing. and with SAM is example, after it is added into substrate, SAM has just formed the sample presentation devices surface that fluid sample may contact. and depend on the composition of the specific SAM that uses, the surface of sample presentation devices of the present invention has different character aspect wettable and to the compatibility of analyte in the fluid sample (or it lacks compatibility) aspect. can SAM be added on the sample presentation devices of the present invention by the mode that produces zones of different (regional character has reflected the employed SAM in specific region). other surface modification technology well known by persons skilled in the art also is contained among the present invention.

Sample presentation devices of the present invention is made up of different zones, one of them is being best aspect the delay fluid sample. sample presentation devices of the present invention can also comprise the different zone of wettable, and one of them is being best aspect the high-sensitivity detection of analyte.

The regional kind that can comprise as for this sample presentation devices surface, they mainly are to rely on them that the different wettables of analyzed sample are characterized, conversely, wettable difference has caused again producing is detained or in conjunction with the different zone of analyte ability in the fluid sample. these zones broadly are called " frontier district ", " liquid holding zone " and " analysis area " though. the present invention also considers to comprise the zone of two or more types, but the present invention only needs to exist two types zone. the present invention also comprises more than one zone in each class-for example, sample presentation devices can comprise a plurality of liquid holding zones, wherein each zone is for fluid sample and/or wherein comprise the analyte different in kind. for the ease of the high flux automatics of (for example mass spectrometer) on the multiple analytical instrument or for its compatibility, multiple zone accurately can be located.

" frontier district " relates to and treats analytic sample and be not wettable substantially zone. and compare with other zone, the frontier district has the highest contact angle for sample.

With regard to sample to be analyzed, the wettable of " liquid holding zone " is relatively large (and more relatively poor relatively than analysis area than the frontier district, following). the contact angle that the contact angle of liquid holding zone is lower than the frontier district relatively (and is relatively higher than the contact angle of analysis area, following). originally, liquid holding zone has the contact angle that is equal to or less than the analysis area contact angle, but because chemistry or physical stimulation make liquid holding zone present than the higher contact angle of analysis area contact angle before chemistry or the physical stimulation, this has caused fluid sample to be guided to one than preferred zone, another zone. in addition, liquid holding zone can have two subclass. in a subclass, liquid holding zone is designed to also play the effect that fluid sample is detained in the combination of basic opposing analyte.In second subclass, liquid holding zone is designed to be detained fluid sample but the analyte in the fluid sample is had sizable combination, so therefore since its can catch analyte and be called as " trapping region ". this second subclass also comprises such surface, promptly should have combination basically to analyte in the surface, but should when accepting chemistry or physical stimulation (such as ultra-violet radiation, electric current or heat), become on the surface analyte not combination basically.

Compare with other zone, " analysis area " is the zone of a wettable maximum for sample (and having minimum contact angle). analysis area is designed to resist the combination of analyte. and aspect size, shape and the surface nature analysis area is being optimized so that improve the sensitivity that the goal analysis thing is analyzed.

In the middle of other benefit, because the interregional wettable difference of sample presentation devices of the present invention is though make it compare the fluid sample that can be detained and handle more volume with employed other biochip in sample treatment. the liquid capacity of sample presentation devices of the present invention depends on the size in zone; But for the border circular areas of 3mm diameter, its liquid capacity can be up to about 100 μ L, and can reach about 70 μ L. sample presentation devices at least and can hold the fluid sample of this quantity and not need physical boundary, reservoir or hole.

In another embodiment of sample presentation devices of the present invention, sample presentation devices can be called " target chip " and be abbreviated as Tn, wherein " n " is the figure notation about zones of different quantity on the sample presentation devices surface, and " n " can be to the Any Digit between the infinity from 2. therefore, for example T2 target chip has two zones, T3 target chip has three zones, or the like. the present invention has considered to comprise many sample presentation devices more than 2 or 3 zones, and the present invention is subject to the specific quantity in zone never in any form. along with region quantity increases, general effect approaches a gradient. the sample presentation devices that the target chip is made up of one or more zones that are designed to resist the analyte combination. for example, as for T2 target chip, this sample presentation devices comprises two zones-be frontier district and analysis area. with the region surface of contact liq sample be designed to analyze thing in conjunction with opposing-be that analysis area is that analyte is in conjunction with opposing. during the drying steps before analyzing, the region surface of contact liq sample has limited analyte effectively. as for T3 target chip, this sample presentation devices comprises three zones-be the frontier district, liquid holding zone and analysis area. with the region surface of contact liq sample be designed to analyze thing in conjunction with opposing-be that liquid holding zone and analysis area are that analyte is in conjunction with opposing. during drying steps, the region surface of contact liq sample is concentrated into analysis area with analyte effectively. therefore, sample presentation devices of the present invention can comprise zones of different, and wherein each zone presents the minimum absorption to analyte.

In another embodiment of sample presentation devices of the present invention, sample presentation devices can be called " catching chip " or " catching/concentrate chip " and be abbreviated as Xn, wherein " n " is the figure notation about this sample presentation devices surf zone quantity, and " n " can be to the Any Digit between the infinity from 2. therefore, for example, X2 target chip has two zones, X3 target chip has three zones, or the like. the present invention has considered to comprise many sample presentation devices more than 2 or 3 zones, and the present invention is subject to the specific quantity in zone never in any form. along with the increase of region quantity, general effect approaches a gradient. and to catch chip and catch/concentrate the sample presentation devices that chip is made up of one or more zones that are designed to bound analyte. the part of being responsible for catching analyte generally comprises special finishing, this is the distinctive feature of trapping region. and these finishinges comprise the biological or chemical part with any combination bound analyte of specificity (as monoclonal antibody) or non-specific (as the charged group based on the electrostatic attraction combination) or this kind gravitation. and these finishinges are except having the ability of catching the goal analysis thing, can also be detained the analyte in the fluid sample so that carry out subsequently modification. therefore, for example, the sample presentation devices of the present invention that comprises the trapping region of monoclonal antibody finishing can be in conjunction with the complementary antigen in the fluid sample, and when the remainder of fluid sample being moved to the other parts of apparatus surface by physical transfer or wettable difference, it can be detained this antigen. by add to the sample presentation devices trapping region other compound (enzyme that for example adds cut-out antigen) modify be detained antigen. then, the other parts that modified antigen are transferred to sample presentation devices are further handled, or its moving-out device is used for analyzing by known technology.

For example, catch chip as for X2, sample presentation devices comprises two zones-be frontier district and trapping region. based on the chemistry on trapping region surface or biological property the region surface of contact liq sample is designed to be able to catch analyte-be the trapping region bound analyte. during the drying steps before analyzing, the region surface of contact liq sample has limited analyte effectively. catch/concentrate chip as for X3, sample presentation devices comprises three zones-be the frontier district, trapping region and analysis area. the frontier district is designed to not wettable basically. trapping region is designed to be able to catch and bound analyte. analysis area is designed to analyze thing in conjunction with opposing. before carrying out analysis by one of multiple known analyzing detecting method, analyte is being shifted between trapping region and the analysis area. during the drying steps before analyzing, analyte has been limited on the analysis area surface that comprises fluid sample effectively. and utilize the character on trapping region surface can finish the transfer of fluid sample from the trapping region to the analysis area-that is to say, if the wettable degree of trapping region is lower than the wettable degree of analysis area, then do not need the physical interventions fluid sample will move to analysis area from trapping region yet. alternatively, thereby can make its character can respond chemistry or physical stimulation (heat for example to trapping region design, ultra-violet radiation) changes, cause the wettable degree of trapping region to be lower than analysis area, and therefore cause fluid sample to move to analysis area from trapping region.

In another embodiment of sample presentation devices of the present invention, sample presentation devices can be above-mentioned target chip and the combination of catching chip. in this embodiment, sample presentation devices is by the surface composition with different functionalities. and the sample presentation devices of these types relates to by mechanical means (for example pipetting by pipette) or other method (for example by wettable difference between the zone) fluid sample from a zone-transfer to another zone. and as an example, " catching-shifts-concentrate chip " (is abbreviated as X2-transfer-T3) and is by the X2 chip that comprises two zones (being frontier district and trapping region) and to comprise three zones (be the frontier district, liquid holding zone and analysis area) the sample presentation devices formed of T3 chip. the transfer of analyte (mechanical transfer or alternate manner shift) occurs between the liquid holding zone of the trapping region of X2 chip and T3 chip.

These sample presentation devices relate to more than one " trapping region ", so that its surface can present the binding affinity to one or more analytes. one of the present invention is characterised in that a lip-deep zone moves to that it has the ability of bound analyte one by one another regional process from sample presentation devices at fluid sample, this is easy to a plurality of different fractions of fluid sample are analyzed, do not separate and do not need to use mechanical intervention to carry out physical property. alternatively, the different wettable character of sample presentation devices of the present invention guide fluid sample to the zones of different of installing, in this process, stay the analyte with different trapping region combinations, and therefore be able to handling liquid samples in turn.

More specifically, relating to trapping region can be used for separating before the analyte to fluid sample detects with compound mode with the embodiment of the sample presentation devices of liquid holding zone combination further, concentrate, purifying and these analytes of modification. therefore, for example, fluid sample can be placed on the T2 chip so that the analyte of sample is limited in analysis area. then, these samples are transferred to comprise the frontier district, the X3 chip of trapping region and analysis area. in this example, trapping region is designed to be able to from fluid sample in conjunction with (and therefore removing) lipid part, so that when sample application during in the X3 chip, it shifts to trapping region (it has the wettable of higher degree) from the frontier district, lipid part in the sample is incorporated into the trapping region surface, and remaining sample moves to analysis area (because it has the wettable of top). in this example, fluid sample is limited on the T2 chip, and subsequently lipid is moved on the X3 chip, this with regard to make final sample in the analysis area analysis be obtain concentrating become no lipid with purifying. owing to trapping region can be designed to be able in conjunction with multiple different analyte, and, has purifying on a large scale so can create owing to can also use the multiple arbitrarily combination in these zones, concentrate, the sample presentation devices of separation and modification ability (about one or more analytes).

Fluid sample can be changed from the mechanism that a sample presentation devices is transferred to another sample presentation devices. use above-mentioned example, can mechanically remove (for example pipetting) concentrating sample from T2 by pipette, and place on the X3 sample presentation devices separately. alternatively, the zone that chemistry or physical stimulation (as ultra-violet radiation) change be can respond by a wettable and T2 and X3 sample presentation devices connected, cause this regional wettable to be higher than T2 device analysis district but be lower than X3 device trapping region so that the zone between T2 device analysis district and X3 device trapping region is exposed to ultra-violet radiation following time, therefore the concentrating sample in the T2 sample presentation devices analysis area is transferred to the trapping region of X3 device. and again, utilization exhibiting high surface (having different wettables and different analyte binding characteristics) and configuration thereof can create has purifying on a large scale, concentrate, the sample presentation devices of separation and modification ability (about one or more analytes).

Sample presentation devices of the present invention (in above-mentioned each embodiment) can further provide the different wettables zone with difformity or pattern (a little example is wherein described in the accompanying drawings). for example, in one embodiment, sample presentation devices has the zone of concentric circles form, middle section is an analysis area, analysis area is surrounded by liquid holding zone, and surrounded by the frontier district by liquid holding zone. because the zone can be made by using multiple light pattern technology, and because known light pattern technology provides very big variation for resulting pattern, so exist here can by those skilled in the art design on a large scale may shape, the multiple zone of pattern and configuration. in addition, the multiple character of wettable zones of different makes to produce and analyte can be directed to lip-deep single or multiple assigned address or predetermined position (addressable sites for example, road or zone) sample presentation devices. " addressable " refers to simply in this article: predetermined site, road or zone can be by specifying with the automatic processing device of sample presentation devices collaborative work of the present invention, so that be stranded in the fluid sample of these assigned addresses or analyte can be handled so that measure the goal analysis thing by analytical equipment. in addition, can shift out from sample presentation devices and be used for carrying out subsequent treatment or operation (is for example modified being present in those fluid samples that pre-determine the position or analyte by another kind of sample presentation devices, purifying, concentrate or the like).

Sample presentation devices of the present invention is suitable for handling biological and abiotic fluid sample. and they also are adapted at using in the analyte detection method widely, and wherein said detection method for example (includes but not limited to) other known analysis methods of analyte in mass spectrometry, multiple chromatography method, immunofluorescence spectroscopic methodology and detection and the measurement fluid sample.

Each of above-mentioned variation all is designed to allow maximum flexibility ratio on to the design and use of sample presentation devices, described sample presentation devices has the analyte of presenting that is better than known method and is used to the ability that detects and analyze. therefore, sample presentation devices of the present invention has the ability that analyte is directed to the analysis area that is designed to strengthen the analyte high-sensitivity detection. and therefore, sample presentation devices of the present invention provides improved analyte deposition.

Sample presentation devices of the present invention further comprises can accept and be detained the device that liquid sample volume reaches about 100 μ L and reaches about 70 μ L at least. and sample presentation devices of the present invention can also be used as the sample positioner, and this device guides analyte to be deposited into through measuring less than about 2 square centimeters of (2mm ²) and preferably less than about 1mm ²Surf zone. guide analyte to be deposited into through measuring less than about 1mm ²Surf zone in be beneficial to and improve the analyte deposition, the raising of simultaneous automatic data collection and detection sensitivity two aspects. therefore, sample presentation devices of the present invention provides the surface that all shows huge purposes aspect liquid holdup ability and the analyte controllable deposition. in preferred embodiments, this kind combination of attributes increases detection sensitivity, with known sample holder mutually specific sensitivity increase about 4 times to about more than 100 times.

In one embodiment, sample presentation devices of the present invention is made up of substrate, the surface of wherein said substrate further is made up of three adjacent areas organizing by concentric arrangement, wherein Zhong Yang analysis area is surrounded by liquid holding zone, and wherein liquid holding zone is surrounded by the frontier district. alternatively, sample presentation devices of the present invention can be made up of substrate, wherein the surface is further by arranging three adjacent areas organizing and form by closing on, wherein partial analysis district and partially liq stagnant area are adjacent one another are, and wherein analysis area and liquid holding zone those parts not adjacent to each other are surrounded by common frontier district.

In the embodiment of sample presentation devices of the present invention, the surface of analysis area has preferably less than about 40 °, be more preferably less than about 30 °, and most preferably less than about 20 ° contact angle. the surface of analysis area preferably presents analyte is had minimum compatibility or combination. the surface of liquid holding zone have preferably about 40 ° to about 95 ° of scopes, more preferably about 60 ° to about 95 ° of scopes, the contact angle in 80 ° to 95 ° scopes most preferably, and further preferably presenting analyte is had minimum compatibility and combination. the surface of frontier district has and is preferably greater than about 95 °, more preferably greater than about 105 °, most preferably greater than about 115 ° contact angle, and further preferably present fluid sample had minimum wettable.

In another embodiment of sample presentation devices of the present invention, the contact angle of analysis area hangs down about at least 10 ° than the contact angle of liquid holding zone, preferably about at least 20 °, more preferably about at least 30 °, and most preferably about at least 40 °, wherein the contact angle of liquid holding zone preferably hangs down about at least 10 ° than the contact angle of frontier district, more preferably about at least 15 °, and most preferably about at least 20 °. in the embodiment of sample presentation devices of the present invention, the surface area of liquid holding zone is than preferably about greatly at least 4 times of the surface area of analysis area, more preferably about at least 10 times, and most preferably about at least 50 times, and the surface area of analysis area is preferably less than about 1mm ², more preferably at about 0.2mm ²To about 0.8mm ²Scope in and most preferably at about 0.4mm ²To about 0.6mm ²Scope in.

Sample presentation devices of the present invention can further be made up of substrate, wherein the surface of this substrate can further be made up of (but being not limited to) 1-1536 right analysis area and liquid holding zone, wherein analysis area and liquid holding zone to or be arranged in concentric or be arranged in and face mutually, and wherein analysis area and liquid holding zone are to being surrounded by common frontier district. by many sample presentation devices that analysis area and liquid holding zone are formed preferably to be similar to standard 96-orifice plate, the mode of 384-orifice plate and 1536-orifice plate is configured, so as with porous plate processor and laboratory fluids handling machine people compatibility.

Accompanying drawing is described

Following description only is representational, be to the replenishing of the disclosure of the Invention of illustrating elsewhere, and it does not limit the scope of the invention.

About Fig. 1 a and 1b, it has illustrated sample presentation devices of the present invention, it shows substrate 1, wherein substrate surface further is made up of three adjacent areas organizing by concentric arrangement, wherein Zhong Yang analysis area 2 is surrounded by liquid holding zone 3, and wherein liquid holding zone 3 is surrounded by frontier district 4. the surface of analysis area 2 has preferably less than about 40 °, be more preferably less than about 30 °, and most preferably less than about 20 ° contact angle, and further preferably present analyte had minimum combination. the surface of liquid holding zone 3 present preferably about 40 ° to about 95 ° of scopes, more preferably about 60 ° to about 95 ° of scopes, the contact angle in 80 ° to 95 ° scopes most preferably, and further preferably presenting analyte is had minimum combination. the surface of frontier district 4 presents and is preferably greater than about 95 °, more preferably greater than about 105 °, most preferably greater than about 115 ° contact angle, and further preferably present fluid sample had minimum wettable.

Further about Fig. 1 a and Fig. 1 b, the preferred embodiment of sample presentation devices of the present invention is that the contact angle of wherein analysis area 2 is than low preferably about at least 10 ° of the contact angle of liquid holding zone 3, more preferably about at least 20 °, more preferably about at least 30 °, and most preferably about at least 40 °, wherein the contact angle of liquid holding zone 3 hangs down preferably about at least 10 ° than the contact angle of frontier district 4, more preferably about at least 15 °, and most preferably about at least 20 °, wherein the surface area of liquid holding zone 3 is than greatly preferably about at least 4 times of the surface area of analysis area 2, more preferably about at least 10 times, and most preferably about at least 50 times, and wherein the surface area of analysis area 2 preferably less than about 2mm ², more preferably at about 0.2mm ²To about 1.8mm ²Scope in and most preferably at about 0.4mm ²To about 1.6mm ²Scope in.

About Fig. 2, sample presentation devices of the present invention is made up of substrate 5, wherein the surface is further concentricity to forming by 16 of analysis area 6 and liquid holding zone 7, wherein all analysis areas and liquid holding zone are to being surrounded by common frontier district 8. in this case, target area and liquid holding zone be to being arranged among the 9mm in the heart, and this will allow six such devices are combined into form corresponding to standard 96-orifice plate.

Further about Fig. 2, the preferred embodiment of sample presentation devices of the present invention is that the contact angle of wherein analysis area 6 is than low preferably about at least 10 ° of the contact angle of liquid holding zone 7, more preferably about at least 20 °, more preferably about at least 30 °, and most preferably about at least 40 °, wherein the contact angle of liquid holding zone 7 hangs down preferably about at least 10 ° than the contact angle of frontier district 8, more preferably about at least 15 °, and most preferably about at least 20 °, wherein the surface area of liquid holding zone 7 is than greatly preferably about at least 4 times of the surface area of analysis area 6, more preferably about at least 10 times, and most preferably about at least 50 times, and wherein the surface area of analysis area 6 preferably less than about 2mm ², more preferably at about 0.2mm ²To about 1.8mm ²Scope in and most preferably at about 0.4mm ²To about 1.6mm ²Scope in.

Importantly it may be noted that: analysis area and liquid holding zone be illustrated circle in all needing not to be in shape such as Fig. 1 a. and analysis area and liquid holding zone all can be taked in order to make sample presentation devices have optimal representation for application-specific the various shape that needs. and in addition, importantly it may be noted that: analysis area and liquid holding zone all need not to be such as Fig. 1 a and the illustrated the sort of each other concentric form of Fig. 2. and analysis area and liquid holding zone can be according in order to make sample presentation devices have the needs of optimal representation and carry out correspondingly position arrangement for application-specific.

About Fig. 3, sample presentation devices of the present invention is made up of substrate 9, the surface of described substrate is further by arranging three adjacent areas organizing and form by closing on, wherein the part of the part of analysis area 10 and liquid holding zone 11 is adjacent one another are, wherein analysis area and liquid holding zone those parts not adjacent to each other are surrounded by common frontier district 12. and the surface of analysis area 10 presents preferably less than about 40 °, be more preferably less than about 30 °, and most preferably less than about 20 ° contact angle, and further preferably present analyte had minimal combination. the surface of liquid holding zone 11 present preferably about 40 ° to about 95 ° of scopes, more preferably about 60 ° to about 95 ° of scopes, most preferably at about 80 ° of contact angles to about 95 ° of scopes, and further preferably presenting analyte is had minimal combination. the surface of frontier district 12 presents and is preferably greater than about 95 °, more preferably greater than about 105 °, most preferably greater than about 115 ° contact angle, and further preferably present fluid sample had minimum wettable.

Further about Fig. 3, the preferred embodiment of sample presentation devices of the present invention is that the contact angle of wherein analysis area 10 is than low preferably about at least 10 ° of the contact angle of liquid holding zone 11, more preferably about at least 20 °, more preferably about at least 30 °, and most preferably about at least 40 °, wherein the contact angle of liquid holding zone 11 hangs down preferably about at least 10 ° than the contact angle of frontier district 12, more preferably about at least 15 °, and most preferably about at least 20 °, wherein the surface area of liquid holding zone 11 is than greatly preferably about at least 4 times of the surface area of analysis area 10, more preferably about at least 10 times, and most preferably about at least 50 times, and wherein the surface area of analysis area 10 preferably less than about 1mm ², more preferably at about 0.2mm ²To about 0.8mm ²Scope in and most preferably at about 0.4mm ²To about 0.6mm ²Scope in.

Importantly it may be noted that: analysis area and liquid holding zone be illustrated circle in all needing not to be as Fig. 1 a, Fig. 2 and Fig. 3 in shape. and analysis area and liquid holding zone all can be taked the multiple shape of the needs in order to make sample presentation devices have optimal representation for application-specific.

About Fig. 4 a, sample presentation devices of the present invention is made up of substrate 13, the surface of wherein said substrate further is made up of three adjacent areas organizing by concentric arrangement, wherein Zhong Yang analysis area 14 is surrounded by liquid holding zone 15, and wherein liquid holding zone 15 is surrounded by frontier district 16. about Fig. 4 b, the shape of analysis area 14 (square) is beneficial to the automatic collection of mass spectrometric data, and this is because the grating in its corresponding in size 36 zones.

About Fig. 5, sample presentation devices of the present invention is made up of substrate 17, wherein said substrate is made up of 96 pairs of analysis areas 18 and liquid holding zone 19, all analysis areas and liquid holding zone are to being surrounded by common frontier district 20. in this case, with concentric paired area arrangement in corresponding to the 9mm of standard 96-orifice plate in the heart. liquid holding zone 19 elongated so that the liquid containing ability is maximum and make among every row the distance between adjacent area minimum. the sinuous pattern that covers on preceding two rows of sample presentation devices has shown that the liquid stream by chromatography eluent in automation fraction collection process deposits the route of being described.

Further about Fig. 5, the preferred embodiment of sample presentation devices of the present invention is that the contact angle of wherein analysis area 18 is than low preferably about at least 10 ° of the contact angle of liquid holding zone 19, more preferably about at least 20 °, more preferably about at least 30 °, and most preferably about at least 40 °, wherein the contact angle of liquid holding zone 19 hangs down preferably about at least 10 ° than the contact angle of frontier district 20, more preferably about at least 15 °, and most preferably about at least 20 °, wherein the surface area of liquid holding zone 19 is than greatly preferably about at least 4 times of the surface area of analysis area 18, more preferably about at least 10 times, and most preferably about at least 50 times, and wherein the surface area of analysis area 18 preferably less than about 2mm ², more preferably at about 0.2mm ²To about 1.8mm ²Scope in and most preferably at about 0.4mm ²To about 1.6mm ²Scope in.

The making of sample presentation devices

Other embodiment of the present invention comprises the method that produces or make above-mentioned sample presentation devices. for example, in the embodiment that the surface is made up of one or more self-assembled monolayers (SAM) (depend on the difference of the SAM that uses and form zones of different), sample presentation devices of the present invention can comprise the multiple SAM zone that produces by known light pattern technology. and therefore, the present invention comprises that further the light pattern technology that is used as a kind of method for optimizing produces the method for the sample presentation devices of being made up of SAM.

More generally, the substrate surface of sample presentation devices of the present invention is generally modified or patterning by the known method of those skilled in the art. as an example, the surface of substrate can be modified or patterning by using one or more self-assembled monolayers (SAM), described self-assembled monolayer is modified substrate surface and its exposed surface of sample presentation devices and can be given substrate special chemical property. for special substrate is selected to comprise 1 °, 2 °, 3 ° or 4 ° of compositions then provide unique surface characteristics and character for substrate surface at interior different SAM. particularly, use a plurality of SAM and carry out base patternization, so that it can comprise a plurality of zones of each region surface characteristic and different in kind. the method for patterning SAM is known in the art, and comprises the UV light patternization, the photolithography patterning, little seal, electron beam patterning and reactive ion etching art.

The zone that produces at substrate surface can be an arbitrary shape, circular. in addition, the zone can be continuous or discontinuous with other zone-be the zone can all adjacent one another are or one or more zones and one or more other zone non-conterminous. preferably have in the zone of the substrate surface generation of sample presentation devices and to treat the different a plurality of zones of analytic sample wettable.

As another embodiment of the invention, it provides makes accurately the positioning analysis thing with the method for the sample presentation devices of convenient automatic data collection.

More specifically, the method for patterned surface, selection, the preparation of self-assembled monolayer and other method of finishing of suitable substrate have hereinafter been described. these descriptions only are representational, and it does not limit the scope of the invention.

Patterning can be carried out by one of several method in the surface of sample presentation devices of the present invention, and described method preferably includes (but being not limited to): (1) is from the UV-light patternization of the self-assembled monolayer (SAM) of the lip-deep alkyl hydrosulfide preparation of coinage metals; (2) from the photolithography patterning of the SAM of the lip-deep alkyl hydrosulfide of coinage metals preparation; (3) from little seal of the SAM of the lip-deep alkyl hydrosulfide of coinage metals preparation; (4) the photolithography patterning of the SAM of the preparation of the alkyl silane on silicon or the glass surface; (5) electron beam patterning and (6) reactive ion etching art. preferably, the patterning on sample presentation devices surface or be described in U.S. Patent number 5 by application, 514, UV-light pattern method in 501 realizes or by being described in U.S. Patent number 5,512, little impression method in 131 is realized, these two patents are integrated conduct with reference to alternatively at this paper, and the patterning on sample presentation devices surface can be realized by the photolithography patterning method of describing in the document of also understanding for those skilled in the art.

About Fig. 6 a to 6h, described the SAM that is made up of the alkyl hydrosulfide on the gold is carried out the step-by-step procedure of UV-light patternization. initial, use moistening process and argon plasma etch suitably to clean by uniting such as the suitable substrate 21 of silicon chip (750 μ m). at first the adhesion layer (25-50nm) with chromium or titanium and tungsten (9: 1) is applied to sheet surface, subsequently coated with the film 22 (100-1000nm) of gold. the metal deposition is finished by sputter (evaporation) process, and this process is calibrated the metal deposition (thickness) of time per unit. and course of injection can carry out or uses from the single fritter of thin slice cutting-out and carry out with complete thin slice.

About Fig. 6 b, by substrate is hatched 1-24 hour in the solution that comprises the 0.05-5mM alkyl hydrosulfide that is dissolved in the ethanol first individual layer 23 is assembled on the gold surface. then with the adorned substrate of ethanol washing surface to remove unnecessary alkyl hydrosulfide, and substrate being flowed down drying at nitrogen. first individual layer 23 is by such alkyl hydrosulfide preparation, and promptly its surface that provides presents greater than about 100 ° contact angle and further presents wettable minimum for fluid sample.

About Fig. 6 c, by in the presence of oxygen, being exposed to the adorned substrate in surface carry out light patternization through the ultraviolet light source of first mask 24, so that, produce the sulfonate monomer that gold surface is shown as low compatibility thus with the monomer oxidation in the exposure region. the opening 25 in the mask causes corresponding to the size of liquid holding zone and the generation of shape facility.

About Fig. 6 d and Fig. 6 e, wash gold surface subsequently and remove sulfonate monomer, and provide golden unmodified zone 26. second individual layer 27 to be assembled on the gold surface by substrate is hatched in the solution that comprises the 0.05-5mM alkyl hydrosulfide that is dissolved in ethanol. then the adorned substrate in surface is washed to remove unnecessary alkyl hydrosulfide with ethanol, and flowing down drying at nitrogen. second individual layer 27 is by such alkyl hydrosulfide preparation, promptly the surface that provides of this alkyl hydrosulfide presents about 40 ° of contact angles to about 95 ° of scopes, and presents analyte is had minimal combination.

About Fig. 6 f, by the ultraviolet light source that in the presence of oxygen, is exposed to through second mask 28 further light patternization is carried out in patterned substrate, so that, produced the sulfonate monomer that gold surface is shown as low compatibility thus with the monomer oxidation in the exposure region. the opening 29 in the mask causes corresponding to the size of analysis area and the generation of shape facility.

About Fig. 6 g and Fig. 6 h, wash gold surface subsequently and remove sulfonate monomer, and provide the unmodified zone 30. on the gold by substrate is dissolved in about 0.05 of ethanol and hatches to the solution of about 5mM alkyl hydrosulfide the 3rd individual layer 31 be assembled on the gold surface comprising. then with the adorned substrate in surface with the ethanol washing to remove unnecessary alkyl hydrosulfide, and flowing down drying at nitrogen. the 3rd individual layer 31 is by such alkyl hydrosulfide preparation, promptly the surface that provides of this alkyl hydrosulfide presents less than about 40 ° contact angle, and presents analyte is had minimal combination.

In this way, the step-by-step procedure of the UV-light patternization of the self-assembled monolayer by adopting the alkyl hydrosulfide preparation from the gold prepares sample presentation devices of the present invention. and above-mentioned UV-light pattern method by the self-assembled monolayer of alkyl hydrosulfide preparation on the gold is representational, and the present invention is not limited only to described method.

About Fig. 7 a to Fig. 7 h, described the SAM that is made up of alkyl hydrosulfide on the gold is carried out the step-by-step procedure of photolithography patterning. the suitably suitable substrate 32 of cleaning such as silicon chip, and spray an adhesion layer and a gold thin film 33 (100-1000nm) thereon.

About Fig. 7 b, substrate hatched in the solution that comprises the 0.05-5mM alkyl hydrosulfide that is dissolved in ethanol first individual layer 34 is assembled on the gold surface. then with the adorned substrate in surface with the ethanol washing to remove unnecessary alkyl hydrosulfide and to flow down drying at nitrogen. first individual layer 34 is that the surface that this alkyl hydrosulfide provides presents less than 40 ° contact angle and further presents analyte is had MIN combination by such alkyl hydrosulfide preparation.

About Fig. 7 c, before lithographic printing, in the adorned substrate in surface, being coated with one deck photoresist 35. photoresists and can being and transfer cloudy or transfer sun. negative photoresist causes that photoresist solubility descends in the exposure area, therefore providing negative-appearing image with respect to mask. positive photoresist causes photoresist solubility increase in the exposure area, therefore provide erect image with respect to mask. described the use of positive photoresist herein. can pass through plug-in type (dip-type) method application photoresist, but preferably use the spin coating machine to use. can with manufacturer provide about the suggestion of photoresist thickness and process time as guidance.

About Fig. 7 d, to unite use needed ultraviolet light source time and it carry out light patternization by the adorned substrate in surface being exposed to the specific photoresist that adopts. photomask 36 can prepare with multiple common used material (including but not limited to chromium plating quartz, Mylar, acetate and metal form). and the opening 37 in the mask causes the generation corresponding to the feature of the size of analysis area and shape.

About Fig. 7 e, originally use the commercialization solution-treated substrate special to the employing photoresist, the photoresist of this solution dissolving exposure area, and those zones 38 that are not exposed under the ultraviolet light source are kept insoluble relatively. after removing the photoresist of exposure, employing oxygen plasma or UV/ ozone treatment make the alkyl hydrosulfide monomer oxidation in the exposure area, and then generation shows as the sulfonate monomer of low compatibility to gold surface. subsequently, the washing gold surface is to remove sulfonate monomer and golden regional 39. of unmodified is provided

About Fig. 7 f, substrate hatched in the solution that comprises the 0.05-5mM alkyl hydrosulfide that is dissolved in ethanol second individual layer 40 is assembled on the gold surface. then with substrate with the ethanol washing to remove unnecessary alkyl hydrosulfide, and flowing down drying at nitrogen. second individual layer 40 be by such alkyl hydrosulfide preparation, and promptly the surface that provides of this alkyl hydrosulfide presents from 40 ° to 95 ° the contact angle in the scope and further presents analyte is had MIN combination.

About Fig. 7 g and Fig. 7 h, with one of organic solvent (for example acetone, 1-Methyl-2-Pyrrolidone etc.) of the unexposed photoresist of several known dissolving further washing to remove remaining photoresist 38, and as mentioned above, the substrate with the patterning now be made up of two zoness of different was coated with the new photoresist 41. of last layer before lithographic printing

About Fig. 7 i and Fig. 7 j, as mentioned above, by being exposed under the ultraviolet light source through second photomask 42, the substrate of patterning carry out light patternization. the opening 43 in the mask causes the generation corresponding to liquid holding zone size and shape facility. uses at first adopting the special commercialization solution-treated substrate of photoresist, the photoresist of this solution dissolving exposure area, and those zones 44 that are not exposed to ultraviolet light source are kept insoluble relatively. after removing the photoresist of exposure, adopt oxygen plasma or UV/ ozone treatment with the alkyl hydrosulfide monomer in the oxidation exposure area, produce the sulfonate monomer that gold surface is shown as low compatibility thus. wash gold surface subsequently so that remove sulfonate monomer and golden regional 45. of unmodified is provided

About Fig. 7 k and Fig. 7 l, substrate hatched in the solution that comprises the 0.05-5mM alkyl hydrosulfide that is dissolved in ethanol the 3rd individual layer 46 is assembled on the gold surface. with substrate with the ethanol washing to remove unnecessary alkyl hydrosulfide and to flow down drying at nitrogen. the 3rd individual layer 46 is to prepare with such alkyl hydrosulfide, promptly the surface that provides of this alkyl hydrosulfide presents greater than 100 ° contact angle and further presents fluid sample is had minimum wettable. and last, with one of organic solvent of the unexposed photoresist of several known dissolvings further washing provide the patterned surface of forming by three zoness of different thus to remove remaining photoresist 44.

In this way, employing prepares sample presentation devices of the present invention by the photolithography patterning step-by-step procedure of the SAM that alkyl hydrosulfide on the gold is formed. should be understood that, the patterning described order (analysis area before this, it then is liquid holding zone, then be the frontier district again) be optional, and prove opposite order (frontier district before this, it then is liquid holding zone, then be analysis area again) the same suitable with illustrated order. above-mentioned photolithography patterning process by the self-assembled monolayer of alkyl hydrosulfide preparation on the gold is representational, and the present invention is not limited only to described method.

Many alkyl hydrosulfide monomers are suitable for preparing sample presentation devices of the present invention. and synthetic about the alkyl hydrosulfide monomer, they all have description (Laibinis, P.E. to the assembling of individual layer and they in the classification aspect the surface tension on assemble surface; Palmer, B.J.; Lee, S.-W.; Jennings, G.K. (1998) " The Synthesis of Organothiols and Their Assembly intoMonolayers on Gold ", at Thin Films, the 24th volume (Ulman, A. compile) the 1-41 page or leaf, Academic Press, San Diego, CA, this place integrates as a reference).

Above-mentioned survey article is classified with regard to the surface energy pair end portion relevant with alkyl hydrosulfide SAM on assemble surface. and can provide height wettable surface and the therefore suitable part for preparing the analysis area monomer including but not limited to CO ₂H, B (OH) ₂, PO ₃H ₂, CONH ₂Can both provide contact angle less than about 40 ° surface with each above-mentioned part of the existing report of OH.. generally speaking, provide the part of height wettable surface to form by hydrogen bond receptor, hydrogen bond donor and combination thereof. provide the moderate wettable and so the end portion that is suitable for preparing the liquid holding zone monomer include but not limited to: (60 ° of CN, 10), O ₂CCH ₃(63 °, 11), CO ₂CH ₃(67 °, 10), NHCOCH ₃(68 °, 11), SCOCH ₃(70 °, 11), OCH ₃(74 °, 11), CONHCH ₃(76 °, 11), NHCOCF ₃(77 °, 11) and CO ₂CH ₂CH ₃(89 °, 10). show in the bracket be and assemble relevant contact angle in surface and corresponding alkyl chain length. generally speaking, provide the part of moderate wettable surface to trend towards forming by the degree of functionality (functionality) that participates in dipole-dipole interaction. provide minimum wettable surface and so the end portion that is suitable for preparing the frontier district monomer include but not limited to: O (CH ₂) ₂CH ₃(104 °, 11), O (CH ₂) ₃CH ₃(113 °, 16), NHCO (CF ₂) ₇CF ₃(114.5 °, 2), O (CH ₂) ₄CH ₃(115 °, 16), O (CH ₂) ₅CH ₃(115 °, 16), OCH ₂CF ₂CF ₃(118 °, 11) and (CF ₂) ₅CF ₃(118 °, 2). what show in the bracket is contact angle and the corresponding alkyl chain length relevant with assemble surface. generally speaking, provide the part of minimum wettable surface to trend towards forming by hydrophobic and oleophobic degree of functionality.

Preferably, the target area of sample presentation devices of the present invention and liquid holding zone are assembled the monomer preparation of surface opposing protein by giving. and a large amount of SAM to the preparation of alkyl hydrosulfide from gold have carried out clear and definite feature description with regard to the protein adsorption aspect. and the strongest surface of opposing protein of hitherto reported is to be derived to show as widow's (ethylene oxide) (OCH ₂CH ₂) those the surperficial .Prime and the Whitesides of unit monomer at first described these surperficial purposes (Prime, K.L. and Whitesides, G.M.J.Am.Chem.Soc., 1993,115,10714-21, text is quoted as a reference). investigation (Ostuni, the E. of the structure-character relation about the surface of opposing protein adsorption are disclosed; Chapman, R.G.; Holmlin, R.E.; Takayama, S.; Whitesides, G.M.Langmuir, 2001,17,5605-5620, this paper integrates as a reference). recently, confirmed that many amphion SAM show good opposing (Holmlin, the R.E. to protein adsorption; Chen, X.; Chapman, R.G.; Takayama, S.; Whitesides, G.M.Langmuir, 2001,17,2841-50, this paper integrates as a reference), so and since they made up the apparent height wettable and the good opposing of protein adsorption had potential use as analysis area.

In preferred embodiments, the analysis area of sample presentation devices of the present invention is by general formula I: HS (CH ₂) ₁₁-(OCH ₂CH ₂) _mOH (wherein m is between 3 to 7) though monomer preparation. the surface that the monomer of this general formula provides presents about 30 ° of contact angles to about 38 ° of scopes. these surfaces can not present minimum possible contact angle, so but since they the protein bottom line in conjunction with aspect have excellent performance and preferably use them. in addition, preferably with the analysis area monomer of general formula I with can provide contact angle to unite use greater than the liquid holding zone monomer on about 60 ° surface.

Similar and preferably, the liquid holding zone of sample presentation devices of the present invention is by general formula I I:HS (CH ₂) ₁₁-(OCH ₂CH ₂) _mThe preparation of the monomer of R, m=3-7 wherein, and wherein radicals R is to influence surface tension and wettable end portion. preferred but exclusively non-, radicals R is selected from OCH ₃, OCH ₂CN, CO ₂CH ₃, CONHCH ₃And CO ₂CH ₂CH ₃In the part one. each surface that provides in the above-mentioned end part presents about 62 ° of contact angles to about 89 ° of scopes.

Alternative and preferably, the liquid holding zone of this sample presentation devices can be HS (CH by molecular formula ₂) ₁₁OCH ₂C ₆H ₅Monomer preparation. its terminal benzyl moiety (CH ₂C ₆H ₅) sample that is dissolved in organic solvent is had specific use, and its surface that provides presents about 90 ° contact angle.

In preferred embodiments, the frontier district of this sample presentation devices is by fluid sample being had minimum wettable monomer preparation, the wherein water-soluble dissolubility buffer solution of analyte, organic solvent and their mixture. the verified monomer with perfluorinate end portion has specific use (Naud, C. in this; Calas, P.; Blancou, H.; Commeyras, A.J.Fluorine Chem., 2000,104,173-183, this paper integrates as a reference).

The preferred embodiments of the invention are that wherein analysis area is HS (CH by molecular formula ₂) ₁₁(OCH ₂CH ₂) ₃The monomer preparation of OH, liquid holding zone are HS (CH by molecular formula ₂) ₁₁(OCH ₂CH ₂) ₃OCH ₃Monomer preparation and frontier district be HS (CH by molecular formula ₂) ₁₁OCH ₂CH ₂(CF ₂) ₅CF ₃Monomer preparation. these combination of monomers provide surperficial analysis area contact angle, liquid holding zone contact angle and frontier district contact angle to be respectively about 38 °, 62 ° and 117 °.

Another preferred version of the present invention is that wherein analysis area is HS (CH by molecular formula ₂) ₁₁(OCH ₂CH ₂) ₃The monomer preparation of OH, liquid holding zone are HS (CH by molecular formula ₂) ₁₁OCH ₂C ₆H ₅Monomer preparation and frontier district be HS (CH by molecular formula ₂) ₁₁OCH ₂CH ₂(CF ₂) ₅CF ₃Monomer preparation. these combination of monomers provide surperficial analysis area contact angle, liquid holding zone contact angle and frontier district contact angle to be respectively about 38 °, 91 ° and 117 °.

Adopted contact angle and wettable by the next accurate control surface of mixing (binary) self-assembled monolayer of two kinds of alkyl hydrosulfide monomer preparations. (Semal, S.; Bauthier, C.; Vou é, M.; VandenEynde, J.J.; Gouttebaron, R.; De Coninck, J.J Phys.Chem.B, 2000,104,6225-6232, this paper integrates as a reference). mix that by the monomer that will be used for preparing height wettable and moderate wettable surface contact angle is adjusted into scope greater than 40 °. preferably, adopt double base SAM or preparation analysis area or preparation liquid holding zone. alternatively, also can be with ternary and quaternary self-assembled monolayer or preparation analysis area or preparation liquid holding zone. ternary and quaternary SAM are respectively by substituted alkyl mercaptan and the assorted binary mixture that replaces asymmetric alkyl disulfide (that is: HS (CH ₂) ₁₁R ¹And R ²(CH ₂) ₁₁S-S (CH ₂) ₁₁R ³) or two assorted binary mixtures that replace asymmetric alkyl disulfides (that is: R ¹(CH ₂) ₁₁S-S (CH ₂) ₁₁R ²And R ³(CH ₂) ₁₁S-S (CH ₂) ₁₁R ⁴) preparation.

About Fig. 8 a to Fig. 8 l, described the step-by-step procedure of the photolithography patterning of the SAM that forms by alkyl silane on the silicon. by describing to some extent in the literature with the modification of alkyl dimethyl chlorosilane, alkyl dimethyl alkoxy silane, alkyl three halosilanes or alkyltrialkoxysilaneand reaction pair silicon or glass and being understood by those skilled in the art.

About Fig. 8 a, remove surface contaminant earlier, subsequently with surface oxidation to produce silanol (Si-OH) part, by the suitably suitable substrate 47 of activation of these steps (as silicon chip, glass flake or the silica deposit metallic substrates on it) so that covalently bound alkyl silane. preferably, simply with oxygen plasma treatment, wash and then can provide average silanol density near 4.9Si-OH/nm with the base of oxygen plasma treatment with oxidizing solution (Piranha solution) ²Activating surface 48.

About Fig. 8 b, behind surface active, first alkyl silane individual layer 49 is assembled to silicon face. accurately finish silanization by solution plating or evaporation. first alkyl silane individual layer 49 is preferably by providing contact angle greater than 100 ° and for the alkyl silane preparation on the minimum surface of fluid sample wettability.

About Fig. 8 c, being coated with last layer photoresist 50. these photoresists or transferring cloudy or transfer sun to the substrate of silanization before the lithographic printing. negative photoresist causes the photoresist solubility reduction the exposure area in, therefore providing negative-appearing image with respect to mask. positive photoresist causes the photoresist solubility in the exposure area to increase, therefore providing erect image with respect to mask. the purposes of positive photoresist is described in Fig. 6. and can use photoresist by the plug-in type process, but preferably use the spin coating machine to use photoresist. manufacturer can be with coaching about the suggestion of photoresist thickness and process time.

About Fig. 8 d, use needed ultraviolet light source down and it carry out light patternization by substrate being exposed to unite with the specific photoresist of adopt. photomask 51 can prepare with multiple common used material (including but not limited to chromium plating quartz, Mylar, acetate and metal form). and the opening 52 in the mask causes the generation of the big or small and shape facility of corresponding liquid holding zone

About Fig. 8 e and Fig. 8 f, use the commercialization solution-treated substrate special at first to the employing photoresist, the photoresist of this solution dissolving exposure area, and those photoresists that are not exposed to the zone 53 of ultraviolet light source are kept insoluble relatively. after removing the photoresist of exposure, adopt oxygen plasma treatment to come activating surface 54, prepare further silanization. can accurately realize by solution plating or evaporation at second alkyl silane individual layer 55. silanization of assembling on the silicon face of activation. second alkyl silane individual layer 55 by can provide contact angle about 40 ° to about 90 ° of scopes and for the alkyl silane preparation in conjunction with minimum surface of analyte.

About Fig. 8 g and Fig. 8 h, remove remaining photoresist 53 by further washing substrate with one of several known organic solvents (for example acetone, 1-Methyl-2-Pyrrolidone) that can dissolve unexposed photoresist, as mentioned above, the patterned substrate that will now be made up of two zoness of different before lithographic printing is coated with last layer photoresist 56.

About Fig. 8 i and Fig. 8 j, as mentioned above, by being exposed to through under the ultraviolet light source of photomask 57, with the further light patternization of the substrate of patterning. the opening 58 in the mask causes the generation corresponding to analysis area size and shape facility. then substrate used for the special commercialization solution-treated of employing photoresist, the photoresist of described solution dissolving exposure area, and those photoresists that are not exposed in the ultraviolet source region 59 are kept insoluble relatively. behind the photoresist of removing exposure, adopt oxygen plasma treatment activating surface 60, prepare further silanization.

About Fig. 8 k and Fig. 8 l, the 3rd individual layer 61. of assembling accurately finished silanization by solution plating or evaporation on the silicon face of activation. and the 3rd alkyl silane individual layer 61 is by providing contact angle to be lower than about 40 ° and to the alkyl silane preparation on the surface of analyte bottom line combination. and final, by with one of organic solvent of the unexposed photoresist of several known dissolving further the washing substrate remove remaining photoresist 59, the surface of the patterning of being made up of three zoness of different is provided thus.

In this way, sample presentation devices of the present invention can adopt the step-by-step procedure by the photolithography patterning of the SAM of alkyl silane preparation on the silicon to prepare. it is to be noted, the patterning described order (frontier district before this, it then is liquid holding zone, then be analysis area again) be optional, and verified opposite order (analysis area before this, it then is liquid holding zone, then be the frontier district again) the same suitable with illustrated order. above-mentioned photolithography patterning method by the self-assembled monolayer of alkyl silane preparation on the silicon is representational, and the present invention is not limited only to described method.

Many alkyl silanes are suitable for preparing sample presentation devices of the present invention. and alkyl silane mostly is that commerce can get, and their synthetic and their purposes in finishing are had gained some understanding. (Shriver-Lake, L.C. (1998) " Silane-modified surfaces for biomaterialimmobilization " Immobilized Biomolecules in Analysis:A PracticalApproach (Cass, T. and Ligler, F.S. compile) chapter 1, the Oxford University Press, the Oxford, Britain, this paper integrates as a reference).

Employing and above-outlined be diverse ways slightly, can be with the silicon face apparatus nucleophilic suitable alkyl silane derivatization first time partly of activation, can give required wettable end portion by affix and make further functionalization of this nucleophilic part. alternatively, in the time can obtaining having the alkyl silane of suitable end portion, can modify the surface with a step. the end portion of suitable preparation sample presentation devices of the present invention include, but is not limited to recited above those.

In preferred embodiments, prepare the analysis area of this sample presentation devices at first with the 3-TSL 8330, then further with its functionalization so that general formula III to be provided: (XO) ₃Si-CH ₂CH ₂CH ₂NHCOCH ₂(OCH ₂CH ₂) _nThe immobilization silane of OH, X or be connected in silicon face or be connected on the contiguous immobilization silane wherein, and wherein n between the 4-8. the monomer of general formula III provides contact angle on about 30 ° of surfaces to about 40 ° of scopes. though these surfaces can not present minimum possible contact angle, so but because they preferably use them having excellent performance aspect the bottom line conjugated protein. in addition, preferably with the analysis area monomer of general formula III with can provide contact angle to unite use greater than the liquid holding zone monomer on 60 ° surface.

Similar and preferably, the liquid holding zone of sample presentation devices of the present invention is at first by the preparation of 3-TSL 8330, then further functionalization so that general formula I V:(XO to be provided) ₃SiCH ₂CH ₂CH ₂NHCOCH ₂(OCH ₂CH ₂) _nThe immobilization silane of R ', X or be connected in substrate or be connected on the contiguous monomer wherein, wherein n between 4-8, and wherein R ' for influencing surface tension and wettable end portion. preferred but exclusively non-, radicals R ' be selected from CH ₃, CH ₂CN, CH ₂CO ₂CH ₃, CH ₂CONHCH ₃And CH ₂CO ₂CH ₂CH ₃In the part one. each of above-mentioned end part all provides contact angle on about 60 ° of surfaces to about 90 ° of scopes.

In preferred embodiments, the frontier district of sample presentation devices of the present invention with one step from general formula V is: (CH ₃) ₂(X ') SiCH ₂CH ₂-(CF ₂) ₇CF ₃The preparation of (wherein X ' is the surface reaction part) alkyl silane, wherein said alkyl silane has minimum wettable to water-soluble sample.

Can adopt multiple alternative finishing chemical method and patterned surface method to prepare sample presentation devices of the present invention. recently, patterning aspect for protein opposing surface, the polymerization composition of material causes people's interest. can be by to surface transplantation polymerization composition or form from the teeth outwards to aggregate into and (for example assign to further functionalization by the patterned surface of alkyl hydrosulfide or alkyl silane SAM initial preparation, Husemann, M.; Mecerreyes, D.; Hawker, J.L.; Hedrick, R.S.; Abbott, N.L.Angew.Chem.Int.Ed.1999,38,647-649; Shah, R.R.; Merreceyes, D.; Husemann, M.; Rees, I.; Abbott, N.L.; Hawker, C.J.; Hedrick, J.L.Macromolecules 2000,33,597-605 and Hyun, J. and Chilkoti, A.Macromolecules 2001,34,5644-5652, this paper all integrates as a reference). recently, disclose about carry out first report (Deng, the T. of patterned surface by the absorption of closure copolymer; Ha, Y.-H.; Cheng, J, Y.; Ross, C.A.; Thomas, E.L.Langmuir, 2002,18,6719-6722, this paper integrates as a reference). verified, degree and the SAM that has three (ethylene glycol) base of polymer thin film opposing absorption of proteins that migrates to SAM is quite or than its better (Chapman, R.G.; Ostuni, E.; Liang, M.N.; Meluleni, G.; Kim, E.; Yan, L.; Pier, G.; Warren, H.S.; Whitesides, G.M.Langmuir 2001,17,1225-1233, this paper integrates as a reference).

Be to be understood that, even some part in the fluid sample also still is detained on the surface of wettable minimum, even under non-special mode, also be like this only. in fact, the advantage of sample presentation devices of the present invention has been facilitated on this kind surface by the ability (is not those parts of target molecule by removing) that for example strengthens its concentrating analysis in subsequent analysis. and this is particularly useful under the situation that the abiotic part that possibility interference analysis thing is analyzed is detained.Yet, the surface of this sample presentation devices is not limited only to this example, but can also comprise such surface, promptly outside analysis area the zone in bound fraction, thereby this part can be independent of the sample that comprises analyte handles or processes. really, use sample presentation devices of the present invention can be detained with the part of analytical biochemical method analysis to any, store, transhipment and subsequent analysis. therefore, the present invention can make part be stranded in the zone that has outside the top wettable zone, yet and be worthwhile to the subsequent analysis of these parts., a considerable amount of goal analysis things generally are not stranded in the zone that has outside the wettable zone of top.Therefore, under the example case of analyte being analyzed by the laser desorption spectroscopic methodology, the goal analysis thing that is stranded in maximum wettable zone is not to carry out desorb with the state that combines with the sample presentation devices surface.

The purposes of sample presentation devices and application

Only be representational about the multiple use of sample presentation devices of the present invention and the description of application hereinafter, it does not limit the scope of the invention.

Sample presentation devices of the present invention with multiple analytical technology and program coupling on have a lot of purposes. therefore, the present invention includes the method for using above-mentioned sample presentation devices. more specifically, the present invention includes and use sample presentation devices of the present invention identifying the method that analyte exists and analyzes a plurality of samples in the sample on the sample presentation devices or on a plurality of sample presentation devices.

In fact, sample presentation devices of the present invention can be united use with any method of analyte in detection, evaluation or the measurement fluid sample. and the example of this type of analytical method includes but not limited to that MALDI-MS or this sample presentation devices of electron spray ionisation MS. especially are fit to unite with the high throughput analysis measuring technique, for example use in MALDI-MS, this sample presentation devices analysis area is beneficial to the high flux data acquisition modes and is configured therein.

The analyte that sample presentation devices of the present invention can also be used for the operating liquid sample and wherein be comprised. the different wettable character that can be designed to based on sample presentation devices surface institute with catch character, the sample presentation devices design can be used for operating, concentrate, the location, store, shift (by with do not pass through mechanical intervention), reclaim (by or do not pass through mechanical intervention), analyze, modify or processing (modifying reagent) by operational analysis thing on sample presentation devices, perhaps classification separating liquid sample or the analyte that wherein comprised. in addition, owing to sample presentation devices of the present invention can be designed to be able to by (for example responding chemistry or physical stimulation, heat, ultra-violet radiation, pressure, electromagnetic radiation) realizes any function in these functions, so sample presentation devices of the present invention can reversible or irreversibly realize these functions, can also further realize the multiple combination of these functions by response external force.

In fact, any fluid sample (with analyte) can both be connected use with sample presentation devices of the present invention. for example, the present invention can be used in the fraction that analysis is reclaimed from liquid chromatography. and the present invention can be used in analysis from the protein spots of 2D gel electrophoresis cutting-out or the enzymic digestion thing for preparing from the fraction that affinity chromatography (being ICAT) is collected. and the present invention can also be used to analyze the sample that reclaims from surface plasma resonance biosensor. and the present invention can also be used to use standard porous format Robotics and analytic approach to carry out sample transfer in 1: 1. really, sample presentation devices of the present invention can be used for handling and operate the fluid sample that obtains from any source almost, and no matter this sample is laboratory experiment result (for example enzymic digestion thing and the surface plasma resonance biosensor sample example of above determining), still the water sample of river (for example from) that from environment, obtains, still (for example human urine sample) that directly obtains from live organism.

The present invention can also be used for sample and store so that file or further analysis. in other words, promptly after being transferred to analysis area, fluid sample do not need immediately analyte that fluid sample comprises to be detected and analyzes.

Therefore, multiple embodiments of the present invention provides the sample presentation devices of exercising multiple liquid handling function, and described function includes but not limited to sample/analyte processing and liquid deposition, delay, transfer, locatees and relocate and store. and this paper provides sample presentation devices of the present invention these more multi-purpose examples.

About Fig. 9 a to Fig. 9 f, set forth a plurality of steps in the sample drying process. the cross-sectional view strength of sample presentation devices of the present invention has shown the surface that is deposited in the substrate 62, this surface is made up of three different zones, wherein Zhong Yang analysis area 63 is surrounded by liquid holding zone 64, and wherein liquid holding zone 64 is further surrounded by frontier district 65.

About Fig. 9 b, fluid sample is dripped at 66 on the sample presentation devices surface, the volume of original sample drop is confined to the surface of analysis area 63 with liquid holding zone 64 simultaneously. to the restriction of sample drop from the surface tension relevant with the limited wettable of frontier district 65. when deposition, the contact angle of sample drop equals the contact angle that selectivity ground is present in the drop on the liquid holding zone approx.

About Fig. 9 c to Fig. 9 e, when sample drop was dry owing to evaporation, the radius of drop and contact angle all reduced, and arrived the radius that is equivalent to analysis area until droplet radius.

About Fig. 9 f, when the radius of the radius of sample drop 67 and analysis area 63 conforms to, the contact angle that can find sample drop is approximately equal to the contact angle that is present in the drop on the analysis area. when sample drop continues drying owing to evaporation, the radius of sample drop no longer further descends, but keep constant, this moment, analyte was deposited as skim on the analysis area surface. by this mode, when being deposited on the lip-deep water-soluble sample drying of various volumes up to about 100 μ L of sample presentation devices, can provide the skim analyte that is limited in being equivalent in the analysis area zone.

For example, has diameter 3.0mm (the about 7.069mm of surface area ²) liquid holding zone and diameter 0.5mm (the about 0.196mm of surface area ²) sample presentation devices of the present invention of analysis area can be limited in the deposition of analyte than in the little about 36 times analysis area surface area of liquid holding zone surface area, therefore the corresponding increase of average analysis thing surface concentration is about 36 times. and the result, above-mentioned in principle sample drop drying process improves sensitivity about 36 times potentially.

About Figure 10 a to Figure 10 d, when lacking analysis area (only having liquid holding zone 68 and frontier district 69), radius does not significantly reduce during sample drop 70 dryings, cause analyte at most of liquid holding zone surface deposition 71. about Figure 10 e to Figure 10 h, when lacking liquid holding zone (only having analysis area 72 and frontier district 73), the volume of sample drop 74 is subjected to the restriction of the liquid containing ability of analysis area 72. and radius does not significantly reduce during sample drop 74 dryings, causes analyte at most of analysis area surface deposition 75.

In the method described in Fig. 9 b to Fig. 9 f detection sensitivity is significantly increased. reference diagram 9a to Fig. 9 d and Figure 10 a to Figure 10 d can understand this phenomenon better. under the situation that lacks analysis area, (see Figure 10 a), yet the average analysis thing surface concentration of per unit area equals total analyte concentration divided by surface area in the liquid holding zone of describing in Figure 10 a 68., exist under the situation at the analysis area that Fig. 9 a describes, the deposition of analyte is limited in analysis area, wherein per unit area average analysis thing surface concentration equals total analyte concentration divided by the analysis area surface area. therefore, the existence of the analysis area of being described among Fig. 9 a 63 makes the average area concentration of analyte increase, the increase of concentration equals the ratio of the surface area of the analysis area of describing 63 among the surface area of the liquid holding zone of describing 68 among Figure 10 a and Fig. 9 a. because the surface area of analysis area is significantly less than the surface area of liquid holding zone, significantly increase so the restriction analysis thing is deposited on the average surface concentration that the surf zone of analysis area just causes presenting to mass spectrometric analyte, detection sensitivity correspondingly increases thereupon.

The video contact angle image that shows among Figure 11 a to Figure 11 h has confirmed the analyte restriction character of analysis area, this character increases detection sensitivity. about Figure 11 a, prepare sample presentation devices of the present invention with liquid holding zone that is measured as about 1.6mm OD and the analysis area that is measured as about 0.7mm OD. in order to be easy to observe focusing effect, analysis area departs from the center and places. after a water is applied to biochip surface, can observe the surface area that it promptly is limited in self corresponding liquid holding zone and analysis area. write down initial left side and right side contact angle, find that two contact angles all are 57.1 °, this numerical value is corresponding to the numerical value that the surface showed by the proprietary preparation of liquid holding zone monomer. in the process of drop drying (seeing Figure 11 b to Figure 11 h) owing to evaporation, observed radius and contact angle all reduce, be equivalent to the analysis area radius until droplet radius. in addition, in the droplet drying process, can observe drop centered moves right so that allow drop that himself is concentrated on analysis area. in Figure 11 h, write down left side and right side contact angle, find that they all are 35.4 °, this numerical value is corresponding to the numerical value that the surface showed by the proprietary preparation of analysis area monomer. and in Figure 12, draw and summed up the drop height that writes down simultaneously with Figure 11 a to Figure 11 h institute rendering image collection, width and contact angle data.

Figure 13 has confirmed the liquid containing ability that liquid holding zone is outstanding. the photo of 16-of the present invention site sample presentation devices has shown that volume is the delay of the sample drop of 5 μ L-70 μ L. and right relative to being the unique factor that significantly limits the sample drop volume of adjacent analysis area and liquid holding zone.

Figure 14 a and Figure 14 b have further confirmed the analyte restriction character of analysis area. (Figure 14 is the photo of 16-of the present invention site sample presentation devices a) to first photo, wherein depositing the sample drop of volume between the 5 μ L-40 μ L on 8 site surface in 16 sites. each sample drop all comprises the soluble dye of equal amount. and second photo (Figure 14 b) is that same sample presentation devices is at the dried photo of sample drop. near the present biochip surface of deposition of dye analysis area. in order to compare, the relative size of analysis area and liquid holding zone is added on the biochip. in this case, need excessive dyestuff so that visible material can be provided, it causes strong-focusing to analyze the shortage of spot.

Can adopt sample presentation devices of the present invention to promote high sensitivity Mass Spectrometer Method to following chemistry and biological analyte, described analyte is selected from but is not limited to: large biological molecule such as peptide, protein, enzyme, zymolyte, zymolyte analog, enzyme inhibitor, polynucleotides, oligonucleotides, nucleic acid, carbohydrate, oligosaccharides, polysaccharide, avidin, streptavidin, agglutinin, pepsin inhibitor, protease inhibitors, A albumen, agglutinin, heparin, G albumen, concanavalin; The fragment of above-mentioned large biological molecule such as nucleic acid fragment, fragments of peptides and protein fragments; Above-mentioned large biological molecule compound such as nucleic acid complexes, protein-DNA compound, genetic transcription compound, gene translation compound, film, liposome, membrane receptor, receptors ligand compound, signal transduction path compound, enzyme-substrate, enzyme inhibitor, peptide complexes, protein complex, carbohydrate complexes and polysaccharide compound; And little biomolecule is such as amino acid, nucleotides, nucleosides, sugar, steroids, lipid, metal ion, medicine, hormone, acid amides, amine, carboxylic acid, vitamin and coenzyme, alcohol, aldehyde, ketone, aliphatic acid, porphyrin, carotenoid, plant growth regulator, phosphate and nucleoside diphosphate sugar, synthesized micromolecule such as medicinal or treat effective agent, monomer, peptide analogues, the steroids analog, inhibitor, mutagens, carcinogen, anti-mitosis medicine, antibiotic, ionophore, antimetabolite, amino acid analogue, antiseptic, transport inhibitors, surface-active agents (surfactant), contain the amine combinatorial libraries, dyestuff, toxin, biotin, the biotinylation compound, DNA, RNA, lysine, n acetylglucosamine n, Procion is red, glutathione, adenylic acid, mitochondria and chloroplaset depressant of functions, electron donor, carrier and acceptor, synthetic protease substrate and analog, phosphatase substrate and analog, esterase and lipase substrate and analog and protein modification reagent. in addition, the analyte of handling by sample presentation devices of the present invention can be an abitotic substance, it includes but not limited to that the polymer that synthesizes such as oligomer and copolymer are as poly-alkylene, polyamide, poly-(methyl) acrylate, polysulfones, polystyrene, polyethers, polyvinylether, polyvinyl ester, Merlon, polyvinylhalide, polysiloxanes and top any two or more copolymer, and other is such as the abiotic analyte of pesticide.

Can be with the water-soluble dissolubility buffer solution of analyte, in organic solvent or their mixture. buffer solution preferably is selected from those buffer solutions by the volatile component preparation, described volatile component includes but not limited to: ammonium acetate, carbonic hydroammonium, ammonium carbonate, ammonium citrate, second triethylenetetraminehexaacetic acid ammonium and carbon triethylenetetraminehexaacetic acid ammonium, triethylammonium formate, the acetate trimethylammonium, carbonic acid trimethylammonium and formic acid trimethylammonium. because the existence of detergent can be offset the analyte restriction character of analysis area, so comprising the water-soluble sample of the non-volatile detergent of high concentration (＞0.1%) will desalination before analysis. organic solvent preferably is selected from those known organic solvents that easily mixes and promote the biological analyte dissolving with water-soluble buffer solution, described organic solvent includes but not limited to: acetate, acetone, acetonitrile, ethanol, N, dinethylformamide (DMF), N, N-dimethyl sulfoxide (DMSO) (DMSO), formic acid, seven fluoro butyric acid, methyl alcohol, N-methyl pyrrolidone (NMP), 2,2,2-trifluoroethanol and trifluoroacetic acid.

In sample drying process, can the heated sample presentation devices (perhaps be placed on the surface of heat block or place under the infrared lamp or place under the thermal current) so that promote the evaporation of high boiling organic solvent or just shorten the needed time of sample drying simply.

Laser desorption flying time mass spectrum analysis method (it is to use a kind of preferred analytical method of sample presentation devices Measurement and analysis thing of the present invention) need be at a kind of material of sample presentation devices surface applications (matrix) so that absorb energy and then the ionization of assistant analysis thing. and usually the reagent as biological analyte detection matrix comprises trans-3,5-dimethoxy-4 '-hydroxycinnamic acid (sinapic acid, SA), alpha-cyano-4-hydroxycinnamic acid (HCCA) and 2,5-dihydroxy-benzoic acid (DHBA). because the solubility of above-mentioned matrix in water is limited, so the mother liquor of these reagent comprises the organic solvent of 50%-100% usually. when uniting use with sample presentation devices of the present invention, need join in the water-soluble sample at the mother liquor that sample application will be comprised matrix before the sample presentation devices. alternatively, the mother liquor that can will comprise matrix after sample deposition and drying is applied to the sample presentation devices surface. and in this case, the preferred use comprises the mother liquor of high percent organic solvent so that the dissolving of analyte in mother liquor that is deposited on the analysis area surface minimized.

Sample presentation devices of the present invention has many application. and the example that can be used in sample type of the present invention includes, but is not limited to not carry out any processing before analysis and sample of directly analyzing and the indirect analysis sample of analyzing again after some type processing.

The sample type example that can use and belong to the direct analysis classes of not carrying out any processing before analysis in the present invention includes, but is not limited to biofluid; Tissue and cell extract and fraction, cell, bacterium, virus; Culture medium; Environment liquid; The surrounding air sampling; Surrounding medium extract (soil extract thing, solid waste extract, wiping eluate, air filtration eluate); Forensic samples; And storehouse (combinatorial chemical library, oligonucleotide library, peptide storehouse, carbohydrate storehouse, lipid storehouse, cell and composition storehouse; Chromosome storehouse and virus base and other larger protein and nucleoprotein assemblage storehouse).

The sample type example that can use and belong to the indirect analysis class processing post analysis of some types (promptly) in the present invention includes, but is not limited to LC (LC) effluent; Gas chromatography (GC) effluent; The gel eluate; The sample digestion of LC effluent or gel eluate; The mass spectrum effluent; The eluate of surface plasma body resonant vibration (SPR) or other biology sensor; The desalting column effluent; The solid phase extractions effluent; The environmental sample of liquid phase fractionation; The sample of deriving of above-mentioned any sample; And other chemistry or physical treatment and any combination thereof.

Sample presentation devices of the present invention further is beneficial to from the mass spectral analysis of the biological analyte of classification separation scheme (perhaps adopt the post LC or adopt electrophoresis) recovery. particularly, its effectiveness is derived from the liquid containing ability of device, and (this makes it possible to dwindle without sample volume in advance and directly collects the chromatography fraction, the electrophoresis purification of samples, sample that reclaims from sample presentation devices and the sample that reclaims from biology sensor) and the combination of the raising (this makes it possible to automatic data collection) of the accurate location of sample and detection sensitivity. the liquid containing ability that sample presentation devices of the present invention provides makes it possible to directly collection from following technology (but being not limited to following technology): affinity chromatography, hydrophobic interaction chromatography, ion-exchange chromatography, the fraction that immobilized metal ion affinity chromatography and size exclusion chromatography reclaim and from relating to the fraction that the quadrature that utilizes two or more cited chromatography methods continuously separates recovery. in addition, sample presentation devices is in the 96-hole of standard, validity in 384-hole and the 1536-hole format makes collects and be processed into possibility on porous plate processing unit (plant) and laboratory fluids handling machine people based on the sample of biochip. therefore, can adopt this sample presentation devices to make high flux mass spectral analysis platform become possibility, in order to support the appearance of proteomics and other key areas of chemistry and biotechnology, need this high flux mass spectral analysis platform.

The identification of proteins in the present age is usually directed to the protein by post LC purifying or the enzymic digestion thing of the protein that cuts down from 2-dimension running gel. and the protein digestibility thing usually need be with reverse LC (RPLC) or solid phase extractions (SPE) desalination before mass spectral analysis. and the sample presentation devices of present invention is applicable to direct collection and the subsequent analysis to the protein digestibility thing by efficient RPLC or SPE desalination.

As particular example, surface plasma body resonant vibration (SPR) biology sensor adopts immobilized protein to study protein-protein interaction and other biology interacts. unfortunately, reclaim the eluent that analyte needs large volume from biology sensor, and for best mass spectral analysis, the concentration of analyte is too low in the sample. sample presentation devices of the present invention is suitable for directly collecting the analyte that reclaims from bio-sensor system; It can be assembled into standard 96-hole format so that also can automatic collection sample be used for mass spectral analysis and can concentrate the large volume fluid sample with the sample collection device of built-in biometric sensor system is compatible.

The liquid containing limitation that accompanies with known mass spectrograph sample presentation devices has promoted the development of multiple micro-column LC method, described method relate to the small-sized pipette that uses filling that the microchromotography medium is arranged pipette point (as

). it is reported, can in protein digestibility thing desalination processes, be accompanied by the mass spectrograph of technology formerly that micro-column method that sample volume reduces is enough to that sample directly is applied in and is used to be detained sample. sample presentation devices of the present invention is fit to directly collecting and subsequent analysis by the protein digestibility thing of micro-column RPLC desalination.

Usually, for above-mentioned sample, can use sample presentation devices of the present invention to finish following process: to concentrate; Dilution; The location; Transhipment; Store; Analysis is presented; Classification separates; Washing; With use back processing (comprising digestion, derivatization and wash-out). should be appreciated that this inventory and not exhaustive and only provide synoptically sample presentation devices of the present invention the example of the multiple application that can use.

In case sample application has been carried out the operation determined above any on sample presentation devices of the present invention and to fluid sample thereon mobile, so just can carry out following application: MALDI-MS after shifting out on the sample presentation devices or from installing; Other analytical technique of mass spectrum; Surface plasma body resonant vibration (SPR); Fluorescence; AFM (AFM); Spectroscopic methodology; Bioluminescence and chemiluminescence; The x X-ray photoelectron spectroscopy X; The elliptical polarization technology; Electrochemical Detection; Phosphorescence and UV spectroscopic methodology, visible light spectrometry and infrared (IR) spectroscopic methodology. should recognize that this only is the part inventory of this type of application. it should also be understood that, any above-mentioned analysis all can be combination and/or series connection, and can carry out these analyses to analyte directly or indirectly according to suitable situation.

Expect that sample presentation devices of the present invention can be applied to many fields, and these fields comprise and contain but be not limited to such as genomics, proteomics, pharmacogenomics, physiology group (physiomics), poisonous substance group (toxinomics), metabolism group (metabonomics), drug discovery/medicament research and development/clinical testing monitoring, toxicology, diagnostics, environment, biology sensor and biological and chemical weapon/bioterrorism. a few specific embodiments of using this sample presentation devices has hereinafter been described. following description only is representational, and does not limit the scope of the invention.

Genomics: mass spectrometry is applied to genotype and there is individual basic premise the phenotype aspect: promptly before ionization to the nucleic acid analyte desalination. traditionally, before placing on the MALDI source, sample carries out desalination. in one embodiment, the sample presentation devices of X3 format can be finished desalination, while condensed nucleic acid analyte. this embodiment is made up of the analysis area of anti-phase trapping region and the combination of opposing analyte. and another embodiment is made up of X4, wherein used two trapping regions and single analysis area. in concentric arrangement, outside trapping region will be by combining the polynucleotides analyte with the complementary hybridization of fixing capture probe specifically; Above-mentioned desalination function is then exercised in the trapped inside district, and analysis area is presented analyte and is used for detecting. and in these two embodiments, on same chip, carry out desalination for analysis and present and improved flux, make sample loss drop to minimum and reduced cost.

Drug discovery/research and development/clinical testing monitoring: many medicines are only effective to the part crowd.Trastuzumab (Herceptin) is exactly an example of this phenomenon, and it is only effective to about 30% breast cancer patient. and with the Trastuzumab is example, and for drug design, the hereditary basis of sensitivity and protein basis are indispensable; But in most of situation, can not before expensive and very long clinical testing, just the crowd be divided into possible reactor and non-reactor. one of main challenge of explaining this clinical test results is to understand reaction and non-reactive biology and/or chemical fundamentals. so those knowledge can be used for the further refining of determining of target group and medicine self.

A kind of method that addresses this problem is before treatment, in the therapeutic process and treatment back obtains analysis data (profile) (protein for example from patient, carbohydrate, lipid), and these analysis data and result of treatment being associated. several embodiments of this device can be applied to this research. the sample that will obtain from patient (blood for example, urine, tissue) handles with cited one or more preprocess methods (such as the multidimensional LC) in above describing partly, and the material of classified separation that will be by method for preparing is applied to this device and passs mass spectrum analysis so that concentrate and be. alternatively, can be applied to have one or more apparatus of the present invention of known specificity trapping region through the sample of minimum degree processing. be transferred to the complementary specific trapping region of tool then earlier with analyte or before being transferred to analysis area, perhaps directly be transferred to analysis area. in this kind mode, can connect under automated manner and use or parallel the use in the not homospecific surface of tool, the analyte of classification separation is presented to the identification and analysis zone carried out mass spectral analysis.

Mass spectrometry provides analysis data (complete mass spectrum) and can know and identify purpose specific molecular part. then, mass spectrum is collected into database and analysis data and patient reaction is associated by the multifactor optimization instrument. by this mode, people can find the pattern in analysis data and/or the special molecular entity, its can predicted treatment reaction, monitor treatment reaction and identify and influence the molecular entity of therapeutic response, thereby can carry out the drug design of complexity day by day.

The same with other research field as herein described, the scientific research in this field depends on a large amount of measurement capabilities to analyte in the liquid solution. and sample presentation devices of the present invention and purposes as herein described have provided a kind of important tool that can be used for further studying.

The environment aspect: the existence of pollutant is global work in the analysis environments sample. the particular problem that this research faces is the diversity of the necessary analytic sample of low concentration and institute of analyte, this is because pollutant may be present in gas, liquid and the solid material. usually, this analysis relates to collections, extracts, derives, classification separates and the detection step.

This device can be applied to the analysis of environmental sample in many ways. and these examples are representative; but it is whole by no means. make the device of apparatus trapping region can be directly from gas or liquid medium collection analysis thing. for example; can the hydrophobic pesticide residues thing in the aqueous solution be caught by hydrophobic surface; it can replace liquid/liquid extracting consuming time and that produce harmful waste. then the material of collecting directly is transferred to analysis area; perhaps before being transferred to analysis area, be transferred to complementary specificity trapping region and carry out the classification separation by connecting or walking abreast; perhaps it being shifted out so that analyze with the cited technology of above-mentioned part one or more from device. mass spectrometry is commonly used to identify the pesticide residues thing; but also can use other technology such as immunoassays. as discussed previously; the present invention can also be used to presenting and/or classification separates material from above listed arbitrary environmental analysis step. and apparatus of the present invention can and be presented the platform that they are analyzed to change form as the deriveding analysis thing. for example, silicyl part and/or acetyl group partly can be added on the pesticide that is fixed on the device so that can clearly identify molecular structure.

Biological and chemical weapon/bioterrorismYet: U.S. government is faced with in military and domestic occasion need be easy to platform and the analytical technology that chemistry and biological reagent detect. the challenge of biological warfare investigation is comprised the sample collection and to nontoxic and poisonous organic distinguishing. current battlefield technology for biologic product is the little molecule that utilizes thermal decomposition that biologic artifact is changed into to be easier to by the MS detection., because it is more more special than at present known method to depend on the technology of peptide biomarker, so consider in advance that to a great extent this kind depends on the technology of peptide biomarker. the individuality test that is used for determine fighting the potential exposure of agent relates to breathalyse or blood extraction technique. as the independent biology sensor of warning device in public places or the purposes in the battlefield cause great interest. all these methods all demonstrate collects using Robotics or other remote control thereof to carry out sample, preliminary treatment and present the challenge of sample aspect to detector. can store, operation, concentrating or the technology of purification of samples or those can have potential attraction to defence mechanism to the technology of aerosol collision Sampling Machine (aerosol impactors) coupling of current use. this device can be to be applied to the investigation that biological warfare/bio-terrorism attacks (bioterror) to the described similar mode of environmental sample that is used for. in addition, the device with conventional trapping region can be designed to be able to be used for collect the purpose microorganism from environment or biologicfluid sample, can handle cell (or virus) closes key label and can present these marks so that detect to discharge.

Embodiment

The following example provides the additional detail of composition, manufacturing and purposes about sample presentation devices of the present invention, but it only is a scope representational and that do not limit the present invention in any way.

Example I

The preparation of 11-(3,3,4,4,5,5,6,6,7,7,8,8,8-13 fluoro octyloxies) 11 carbon-1-alkene (1)

3.0mL 1H packs in brown shell bottle (40mL), 1H, 2H, 2H-perfluor octanol (13.7mmol) and to wherein adding 1.4mL 50% aqueous potassium hydroxide (13.7mmol). solution is heated to 80 ℃, stirred 30 minutes and add 3.3mL 11-bromine 11 carbon-1-alkene (1.5mmol). be reflected at 80 ℃ and carried out 52 hours, analyze (hexane) until TLC and show that raw material exhausts. product is cooled to room temperature, add 100mL ethyl acetate and water (2 * 50mL) and salt solution (1 * 50mL) extracting. the ethyl acetate extract passes through dried over mgso, filtering also and obtain the oily residue behind the evaporating solvent in a vacuum. residue is purifying (50 * 300mm on the quick post of silica gel, 0% ethyl acetate/hexane before this, be 10% ethyl acetate/hexane then). the fraction that will comprise the purpose product merges, and obtains the water white oil of 4.52g (64%) formula 1 compound behind the evaporating solvent. ¹H NMR (400MHz, CDCl ₃): δ 5.80 (m, 1H), 4.95 (m, 2H), 3.69 (t, J=6.8Hz, 2H), 3.43 (t, J=6.8Hz, 2H), 2.39 (m, 2H), 2.03 (m, 2H), 1.55 (m, 2H), 1.36 (m, 2H), 1.27 (broad peak m, 10H).

Example II

Thioacetic acid S-[11-(3,3,4,4,5,5,6,6,7,7,8,8,8-13 fluoro octyloxies) undecyl] preparation of ester (2)

1.0g formula 1 compound (1.9mmol) of under argon gas, packing in the dry round-bottomed flask (100mL), and the dried methyl alcohol of adding 10mL. in gained solution, add 426mL thiol-acetic acid (6.0mmol), add 52mg 2 then, 2 '-azo (2-methyl-prop amidine) dihydrochloride (0.2mmol). reaction system hides with paper tinsel system tent, and be exposed under the low pressure mercury lamp light after .4 hour, TCL analyzes (5% ethyl acetate/hexane) and shows that raw material exhausts. and vacuum evaporating solvent obtains the oily residue. and residue is purifying (40 * 300mm on the quick post of silica gel, 0% ethyl acetate/hexane before this, be 5% ethyl acetate/hexane then). the fraction that will comprise the purpose product merges, and obtains the water white oil of 856mg (76%) formula 2 compounds behind the evaporating solvent. ¹H NMR (400MHz, CDCl ₃): δ 3.69 (t, J=6.8Hz, 2H), 3.43 (t, J=6.8Hz, 2H), 2.39 (m, 2H), 2.31 (s, 3H), 1.55 (m, 2H), 1.33 (m, 2H), 1.25 (broad peak m, 10H).

EXAMPLE III

The preparation of 11-(3,3,4,4,5,5,6,6,7,7,8,8,8-13 fluoro octyloxies) hendecane-1-mercaptan (3)

A tool Teflon (Teflon) line silicon diaphragm is installed in brown shell bottle (20mL), the 850mg formula of packing into 2 compounds (1.1mmol), and add 5mL 3N methyl alcohol hydrogen chloride (15mmol). gained solution is heated to 40 ℃ and kept 4 hours. remove the water white oil that solvent obtains 782mg (98%) formula 3 compounds. ¹H NMR (400MHz, CDCl ₃): δ 3.69 (t, J=6.8Hz, 2H), 3.43 (t, J=6.6Hz, 2H), 2.51 (dd, J=7.3,7.6Hz, 2H), 2.39 (m, 2H), 1.58 (m, 4H), 1.32 (t, J=8.0Hz, 1H), 1.25 (broad peak m, 12H).

EXAMPLE IV

11-{2-[2-(2-methoxy ethoxy) ethyoxyl] ethyoxyl } preparation of 11 carbon-1-alkene (4)

In round-bottomed flask (200mL), pack into 27.4mL triethylene glycol monomethyl ester (171mmol) and 9.1mL 50% aqueous NaOH (114mmol). pale yellow solution is heated to 80 ℃, stirred 30 minutes, and dropwise add 26.6mL 11-bromine 11 carbon-1-alkene (114mmol). 80 ℃ of reactions 7.5 hours down, analyze (100% ethyl acetate) until TLC and show that raw material exhausts. product is cooled to room temperature, with 50mL water dilution and with the hexane extracting (3 * 50mL). merge the hexane extract, pass through dried over mgso, obtain the clarification of 20g (56%) formula 4 compounds behind filtration and the vacuum evaporating solvent, water white oil. ¹H NMR (400MHz, CDCl ₃): δ 5.81 (m, 1H), 4.96 (m, 2H), 3.68-3.56 (m, 12H), 3.44 (t, J=6.8Hz, 2H), 3.38 (s, 3H), 2.04 (m, 2H), 1.57 (m, 2H), 1.36 (m, 2H), 1.27 (broad peak s, 10H).

EXAMPLE V

The preparation of thioacetic acid S-(11-{2-[2-(2-methoxy ethoxy) ethyoxyl] ethyoxyl } undecyl) ester (5)

5.0g formula 4 compounds (15.8mmol) of under argon gas, packing in the dry round-bottomed flask (200mL), and the dried methyl alcohol of adding 10mL. to wherein adding 3.6mL thiol-acetic acid (50mmol), add 434mg 2 then, 2 '-azo (2-methyl-prop amidine) dihydrochloride (1.6mmol). reaction system hides with paper tinsel system tent, and be exposed under the light of low pressure mercury lamp after .15.5 hour, TCL analyzes (ethyl acetate/hexane 1: 3) and shows that raw material exhausts. and vacuum evaporating solvent obtains having the residue of strong sulphur sample smell. and residue is purifying (40 * 300mm on the quick post of silica gel, 30% ethyl acetate/hexane, with 50% ethyl acetate/hexane). the fraction that will comprise the purpose product merges, and evaporating solvent obtains the water white oil of 5.83g (94%) formula 5 compounds. ¹H NMR (400MHz, CDCl ₃): δ 3.67-3.54 (m, 12H), 3.44 (t, J=7.2Hz, 2H), 3.38 (s, 3H), 2.86 (t, J=7.2Hz, 2H), 2.32 (s, 3H), 1.57 (m, 4H), 1.36-1.26 (broad peak m, 14H).

Example VI

11-{2-[2-(2-methoxy ethoxy) ethyoxyl] ethyoxyl } preparation of hendecane-1-mercaptan (6)

Not human relations line silicon diaphragm of tool Teflon is installed in brown shell bottle (20mL), and the 5.0g formula of packing into 5 compounds (12.7mmol) also add 7mL 3N methyl alcohol hydrogen chloride (21mmol). gained solution is heated to 40 ℃ and kept 6 hours. and vacuum evaporating solvent obtains the colourless wax gel of 4.40g (98%) formula 6 compounds then. ¹H NMR (400MHz, CDCl ₃): δ 3.67-3.54 (m, 12H), 3.44 (t, J=6.8Hz, 2H), 3.37 (s, 3H), 2.51 (dd, J=7.3,8.0Hz, 2H), 1.57 (m, 4H), 1.32 (t, J=7.6Hz, 1H), 1.26 (broad peak m, 14H).

Example VII A

2-[2-(2-11 carbon-10-thiazolinyl oxygen base oxethyl) ethyoxyl] preparation of ethanol (7)

In round-bottomed flask (250mL), pack into the triethylene glycol (0.5mol) of 67.0mL, and adding 8.0mL50% aqueous NaOH (8mL, 0.1mol). hot solution is heated to 100 ℃, stirred 30 minutes and dropwise added 22.0mL 11-bromine 11 carbon-1-alkene (0.1mol) and obtain producing the deep yellow solution that sodium bromide precipitates. react on 100 ℃ and kept 2.5 hours, analyze (methanol/ethyl acetate/hexane 1: 1: 8) until TLC and show that raw material exhausts. reactant is cooled to room temperature, with 300mL water dilution and with the hexane extracting (3 * 100mL). merge organic extract, with salt water washing (50mL), through dried over mgso and filtration. vacuum evaporating solvent obtains the oily residue. and residue is purifying (50 * 400mm on the quick post of silica gel, methanol/ethyl acetate/hexane 5: 5: 90). the fraction that will comprise the purpose product merges, and obtains the clarified oil of 20.8g (69%) formula 7 compounds behind the evaporating solvent. ¹H NMR (400MHz, CDCl ₃): δ 5.78 (m, 1H), 4.93 (m, 2H), 3.72-3.55 (m, 12H), 3.42 (t, J=7.2Hz, 2H), 2.64 (t, J=5.6Hz, 1H), 2.01 (m, 2H), 1.54 (m, 2H), 1.34 (m, 2H), 1.25 (broad peak s, 10H).

Example VII A I

The preparation of thioacetic acid S-(11-{2-[2-(2-hydroxy ethoxy) ethyoxyl] ethyoxyl } undecyl) ester (8)

2.0g formula 7 compounds (6.6mmol) of under argon gas, packing in the dry round-bottomed flask (100mL), and the dried methyl alcohol of adding 10mL. to wherein adding 2.85mL thiol-acetic acid (40mmol), add 271mg 2 then, 2 '-azo (2-methyl-prop amidine) dihydrochloride (1.0mmol). reaction system is hidden with paper tinsel system tent, and be exposed under the low pressure mercury lamp light after .6 hour, TCL analyzes (methanol/ethyl acetate/hexane 1: 1: 8) and shows that raw material exhausts. and vacuum evaporating solvent obtains yellow oil. and this oil is purifying (50 * 300mm on the quick post of silica gel, methanol/ethyl acetate/hexane 1: 1: 8). the fraction that will comprise the purpose product merges, and evaporating solvent obtains the light yellow oil of 2.44g (98%) formula 8 compounds. ¹HNMR (400MHz, CDCl ₃): δ 3.71-3.54 (m, 12H), 3.42 (t, J=6.6Hz, 2H), 2.83 (t, J=7.2Hz, 2H), 2.66 (broad peak s, 1H), 2.29 (s, 3H), 1.52 (m, 4H), 1.36-1.23 (broad peak m, 14H).

Example I X

2-{2-[2-(11-sulfydryl hendecane oxygen base) ethyoxyl] ethyoxyl } preparation of ethanol (9)

Tool Teflon line silicon diaphragm is installed in brown shell bottle (20mL), and the 2.40g formula of packing into 8 compounds (6.4mmol) also add 5.0mL 3N methyl alcohol hydrogen chloride (15mmol). gained solution is heated to 40 ℃ and kept 4 hours. and vacuum evaporating solvent obtains the colourless wax gel of 2.05g (95%) formula 9 compounds then. ¹H NMR (400MHz, CDCl ₃): δ 3.72-3.55 (m, 12H), 3.43 (t, J=6.8Hz, 2H), 2.71 (broad peak s, 1H), 2.50 (dd, J=7.6,7.4Hz, 2H), 1.62-1.52 (m, 4H), 1.31 (t, J=7.6Hz, 1H), 1.26 (broad peak m, 14H).

Embodiment X

The preparation of 11 carbon-10-thiazolinyl-methylol benzene (10)

Under argon gas, in dry round-bottomed flask (100mL), pack into 5.0g 11 carbon-10-alkene-1-alcohol (29.4mmol) and add 25mL and do N, dinethylformamide. gained solution is cooled to 0 ℃ and add 60% sodium hydride (45nmol) that is dissolved in mineral oil of 2.16g with a part. foaming mixture was stirred 30 minutes in 0 ℃ under argon gas. to cooling, dropwise add in the solution after the stirring and be dissolved in the dried N of 5mL, the 7.7g toluene bromide (45mmol) of dinethylformamide also stirs reactant 3 hours, rise to room temperature simultaneously. by slow adding 100mL ethyl acetate cessation reaction, and with 1N hydrochloric acid (2 * 50mL) and salt solution (1 * 50mL) carries out extracting. with organic layer through dried over mgso, filtering also, evaporating solvent obtains oily residue (9.5g). and residue is purifying (50 * 300mm on the quick post of silica gel, hexane/toluene/acetyl triethyl of 94: 5: 1) and the fraction that will comprise the purpose product merge. last, vacuum evaporating solvent obtains the water white oil of 7.1g (93%) formula 10 compounds. ¹H NMR (400MHz, CDCl ₃): δ 7.32 (d, 4H), 7.28 (m, 1H), 5.81 (m, 1H), 4.95 (m, 2H), 4.49 (s, 2H), 3.46 (t, 2H), 2.03 (m, 2H), 1.61 (m, 2H), 1.35 (broad peak m, 4H), 1.24 (broad peak s, 10H).

Embodiment XI

The preparation of thioacetic acid S-(11 benzyloxy undecyl) ester (11)

5.0g formula 10 compounds (19.2mmol) and 0.520g 2 pack in the light reaction container (250mL) of tool chuck, 2 '-azo ((2-methyl-prop amidine) dihydrochloride (1.92mmol). with seal of vessel, vacuumize and with argon gas recoil (several cycles). in argon gas following time, in reaction vessel, inject 60mL absolute methanol and 0.520g thiol-acetic acid (92mmol) and stirred vessel content. once more container is vacuumized and with argon gas recoil (several cycles). activate the UV lamp and also under argon gas, shine mixture, continue simultaneously to stir 3 hours. constantly cool off (water leg) reactant and in light reaction procedure, temperature maintenance is being lower than 38 ℃. reaction vessel is cooled to room temperature and evaporating solvent and obtains light yellow oil (10.8g). oil purifying (50 * 300mm on the quick post of silica gel, hexane/acetyl triethyl of 98: 2) and the fraction that will comprise the purpose product merge. last, vacuum is removed the water white oil that solvent obtains 5.0g (77%) formula 11 compounds. ¹HNMR (400MHz, CDCl ₃): δ 7.32 (d, 4H), 7.28 (m, 1H), 4.49 (s, 2H), 3.46 (t, 2H), 2.86 (t, 2H), 2.31 (s, 3H), 1.50-1.66 (m, 4H), 1.20-1.40 (broad peak m, 14H).

Embodiment XII

The preparation of 11-benzyloxy hendecane-1-mercaptan (12)

Tool Teflon line silicon diaphragm is installed in brown shell bottle (40mL), the 3.04g formula of packing into earlier 11 compounds (9.03mmol), pack into then 2mL carrene, 1mL hexane and 12mL 4.9N ethanolic hydrogen chloride. gained solution is heated to 40 ℃ and kept 4.5 hours. vacuum evaporating solvent obtains colorless oil residue (2.8g) then. and residue purifying (25 * 450mm, 9: 1 hexane/chloroform) and fraction that will comprise the purpose product on the quick post of silica gel merge. obtain the water white oil of 2.5g (94%) formula 12 compounds behind the vacuum evaporating solvent. ¹H NMR (400MHz, CDCl ₃): δ 7.32 (d, 4H), 7.28 (m, 1H), 4.49 (s, 2H), 3.46 (t, 2H), 2.51 (q, 2H), 1.55-1.65 (m, 4H), 1.20-1.40 (broad peak m, broad peak t, 15H).

Example VII A I

On the silicon base that scribbles gold, prepare self-assembled monolayer

Silicon chip (200mm, P type, one-level silicon 100) cut into independent substrate and it cleaned to provide and have the surface that each substrate is less than 10 particles (0.16 μ m-3000 μ m). in the CPA9900 sputtering system with 5 * 10 ^-7The mm pressure of foundation carries out the metal deposition.In sputtering chamber, clean substrate is also used argon plasma etch, and with The speed sputter become thickness to be

Titanium and tungsten (1: 9) adhesion layer. then with The speed sputter up to

Gold. before shifting out, substrate is cooled off under argon gas stream.

Before the individual layer assembling, with argon plasma under 200W, handle 300 seconds to scribble the gold substrate clean. use the alcohol flushing substrate, (11-(3 then it to be transferred to 0.1mM formula 3 compounds that are dissolved in ethanol, 3,4,4,5,5,6,6,7,7,8,8,8-13 fluoro octyloxies) solution and at room temperature hatching 1-24 hour hendecane-1-mercaptan). last, from assembling is bathed, shifting out the substrate of finishing, flow down drying with 1000 rev/mins of spin rinses and at nitrogen with ethanol. the advancing contact angle that is applied to the suprabasil water droplet of this finishing (0.5 μ L) is between 114 °-120 °. the substrate of finishing is stored in the suitable plastic container that has transparent brown anti-UV lid.

Embodiment XIV

The preparation of patterning sample presentation devices

As mentioned above, the substrate of 24 finishinges of preparation places it in and also uses needle-like alignment (pin-registered) etching stainless steel shadow shield (0.002 inch) that has corresponding to liquid holding zone size and shape facility to cover in the alignment jig customized. and anchor clamps are placed on and are fixed with ratio is 120W/cm ²The transport tape of air cooling ultraviolet (UV) curing systems of low pressure mercury lamp light source on and in 1 hour process through this light source 45 to 75 times. after the UV exposure, substrate is taken out from anchor clamps, flowing down drying with ethanol with 1000 rev/mins of rotation washings and at nitrogen. the substrate that will expose is placed in the formula that is dissolved in ethanol 6 compounds (11-{2-[2-(2-methoxy ethoxy) ethyoxyl] ethyoxyl } hendecane-1-mercaptan) solution of 0.1mM and at room temperature hatched 1 to 24 hour. from bathing, assembling shifts out the finishing substrate of patterning, flow down drying with ethanol with 2400 rev/mins of rotation washings and at nitrogen. the advancing contact angle of water droplet that is applied to liquid holding zone between 60 °-65 °, and when drop is applied to the frontier district its advancing contact angle between 110 °-119 °.

The finishing substrate of patterning is placed in the alignment jig customized, and cover with second needle-like alignment etching stainless steel shadow shield that has corresponding to analysis area size and shape facility. anchor clamps be placed on the transport tape of ultraviolet (UV) curing systems and in 1 hour process through this light source 45 to 75 times. after UV exposes, from anchor clamps, take out substrate, flowing down drying with ethanol with 1000 rev/mins of rotation washings and at nitrogen. the substrate that will expose is placed in 0.1mM formula 9 compounds that are dissolved in ethanol (2-{2-[2-(11-sulfydryl hendecane oxygen base) ethyoxyl] ethyoxyl } ethanol) solution and at room temperature hatched 1 to 24 hour. and last, from anchor clamps, shifting out the finishing substrate of double patterning, flow down drying with 1000 rev/mins of rotation washings and at nitrogen with ethanol. the advancing contact angle of water droplet that is applied to analysis area is less than 47 °. the finishing substrate of double patterning is stored in the suitable plastic container that has brown transparent anti-UV lid.

When above describing multiple embodiments of the present invention, be to be understood that they only occur in the mode of embodiment, do not have limitation. particularly, the physical arrangement of analysis area, liquid holding zone and frontier district is not limited by the foregoing description. and therefore, range of the present invention and scope are not subjected to the restriction of above-mentioned representative embodiment.

Embodiment XV

Sample capacity and location

Be confirmed in the video contact angle image that the analyte restriction character of the analysis area that detection sensitivity increases is shown in Figure 11 a to Figure 11 h.About Figure 11 a, sample presentation devices of the present invention is by liquid holding zone that is measured as about 1.6mm OD and the analysis area preparation that is measured as about 0.7mm OD. for the ease of observing focusing effect, analysis area departs from the center and places. a water is applied to biochip surface, and observe it and self is limited in the surf zone of corresponding liquid holding zone and analysis area rapidly. write down initial left side and right side contact angle, find that two contact angles all are 57.1 °, this numerical value is corresponding to the numerical value that the surface showed by the proprietary preparation of liquid holding zone monomer. in the process of drop drying (seeing Figure 11 b to Figure 11 h) owing to evaporation, observed radius and contact angle all reduce, be equivalent to the analysis area radius until droplet radius. in addition, in the droplet drying process, drop centered moves right so that drop concentrates on analysis area with himself. and find that the left side and the right side contact angle that write down among Figure 11 h all are 35.4 °, this numerical value is corresponding to the numerical value that the surface showed by the proprietary preparation of analysis area monomer. and in Figure 12, draw and summed up the drop height that writes down simultaneously with Figure 11 a to Figure 11 h institute rendering image collection, width and contact angle data.

Embodiment XVI

The liquid containing ability of patterning sample presentation devices

The outstanding liquid containing ability of liquid holding zone is confirmed in Figure 13. and the photo show sample drop retention volume of 16-of the present invention site sample presentation devices is between 5 μ L-70 μ L. and unique factor of significant limitation sample drop volume is adjacent target area and right relative approaching of liquid holding zone.

Embodiment XVII

Analyte guides and concentrates

The analyte restriction character of analysis area further is confirmed in Figure 14 a and Figure 14 b. and (Figure 14 is the photo of 16-of the present invention site sample presentation devices a) to first photo, wherein deposit the sample drop of 5 μ L-40 μ L scope volumes on 8 sites in 16 sites, surface. every dropping liquid drip the HCCA. Figure 14 b that all comprises equal amount be since Figure 14 a describe to carry out drying on the sample presentation devices and concentrated and guide to the photo of the HCCA of analysis area. in order to compare, the relative size of analysis area and liquid holding zone is added on the biochip.

When above describing multiple embodiments of the present invention, it is to be further understood that they only occur in the mode of embodiment, not limitation. particularly, the physical arrangement of analysis area, liquid holding zone and frontier district is not restricted to the described embodiments.Therefore, range of the present invention and scope are not subjected to the restriction of above-mentioned representative embodiment.

Claims

1. the sample presentation devices that is used for the test sample analyte, wherein said sample presentation devices comprises the substrate with surface, wherein the surface is made up of a plurality of different wettables zone, and the zone of analyte is that analyte is in conjunction with opposing basically in the test sample.

2. the sample presentation devices of claim 1, one of wherein different wettables zone is being best aspect the sample delay.

3. the sample presentation devices of claim 1, one of wherein different wettables zone is being best aspect the high-sensitivity detection of analyte.

4. the sample presentation devices of claim 1, wherein substrate is selected from glass, semiconductor, metal, polymer, plastics, has SiO ₂The silicon of layer and have an Al ₂O ₃In the aluminium of layer one or more.

5. the sample presentation devices of claim 1, wherein one or more different wettables zones are made up of self-assembled monolayer.

6. the sample presentation devices of claim 1, it also comprises not wettable basically frontier district and one or more additional areas, and wherein said each additional areas is more wettable than the frontier district.

7. the sample presentation devices of claim 6, wherein one or more additional areas comprise than more wettable liquid holding zone in frontier district and the analysis area more wettable than liquid holding zone.

8. the sample presentation devices of claim 6, wherein the frontier district has than the higher contact angle of each regional contact angle in one or more additional areas.

9. the sample presentation devices of claim 7, wherein the frontier district has the contact angle higher than liquid holding zone contact angle, and wherein liquid holding zone has the contact angle higher than analysis area contact angle.

10. the sample presentation devices of claim 1, but wherein a plurality of different wettables zone comprises not wettable basically frontier district and at least one wet zone of bound analyte basically.

11. be used to store sample presentation devices from the analyte of sample, wherein said sample presentation devices comprises the substrate with surface, wherein the surface is made up of a plurality of different wettables zone, and the zone of wherein storing from the analyte of sample is that analyte is in conjunction with resisting basically.

12. the sample presentation devices of claim 1, wherein sample volume is less than or equal to 100 μ L.

13. the sample presentation devices of claim 1, wherein sample volume is less than or equal to 70 μ L.

14. make the method for the sample presentation devices that is used for the test sample analyte, wherein sample presentation devices comprises the substrate with surface, wherein said method comprises modifies substrate surface producing a plurality of different wettables zone, and wherein in the test sample zone of analyte be that analyte is in conjunction with opposing basically.

15. the method for claim 14, wherein said modification comprise one or more self-assembled monolayers of basad surface applications.

16. the method for claim 15 is wherein used self-assembled monolayer and is comprised and use one or more patterning techniques be selected from UV light patternization, photolithography patterning, little seal, electron beam patterning and the reactive ion etching method with patterned surface.

17. the method for analyte in the test sample, it comprise with sample contact with the sample presentation devices of claim 1 and test sample in analyte.

18. detect the method for analyte in a plurality of samples, it comprises a plurality of samples and the sample presentation devices of claim 1 is contacted and detect analyte in a plurality of samples.

19. the method for claim 17, wherein the method for analyte comprises a kind of method that is selected from mass spectrometry, surface plasma body resonant vibration, fluorescence, AFM, spectroscopic methodology, bioluminescence, chemiluminescence, x X-ray photoelectron spectroscopy X, elliptical polarization technology, Electrochemical Detection, phosphorescence, ultraviolet spectroscopy, visible light spectrometry and the infra-red sepectrometry in the test sample.

20. the method for claim 19, wherein mass spectrometry is the laser desorption ionisation mass spectrometry.

21. use the sample presentation devices of claim 1 to concentrate the method for the sample that comprises analyte, wherein said method is included in concentrating sample in the wettable zone of top.

22. the method for claim 21, wherein the area in top wettable zone is less than 2mm ²

23. the method for claim 21, wherein the area in top wettable zone is less than 1mm ²

24. the method for claim 21, it comprises that also the sample that will concentrate is transferred to one or more additional samples presentation devices in top wettable zone, and wherein each device comprises a plurality of zones different for institute's concentrating sample wettable.

25. the method for analyte in the test sample, it comprises catches the analyte that combines basically with one or more zones of the sample presentation devices of claim 10 from sample.

26. the method for analyte in the test sample, it comprises the material that reduces interfere with subsequent sample treatment process from sample, and wherein said material combines basically with one or more zones of the sample presentation devices of claim 10.

27. store method from the analyte of sample, it is included in and stores analyte on the sample presentation devices, wherein said sample presentation devices comprises the substrate with surface, and wherein the surface is made up of a plurality of different wettables zone and the zone of wherein storing from the analyte of sample is that analyte is in conjunction with resisting basically.

28. handle the method for the sample that comprises analyte, it comprise with sample and the sample presentation devices contact of forming by a plurality of different wettables zone and in top wettable zone concentrating sample, and wherein top wettable zone is that analyte is in conjunction with resisting basically.

29. the method for claim 28, it also comprises the analyte in the sample that detection concentrates in top wettable zone.

30. the method for claim 29, wherein the method for analyte comprises a kind of method that is selected from mass spectrometry, surface plasma body resonant vibration, fluorescence, AFM, spectroscopic methodology, bioluminescence, chemiluminescence, x X-ray photoelectron spectroscopy X, elliptical polarization technology, Electrochemical Detection, phosphorescence, ultraviolet spectroscopy, visible light spectrometry and the infra-red sepectrometry in the test sample.

31. the method for claim 30, wherein mass spectrometry is the laser desorption ionisation mass spectrometry.

32. the method for using the sample presentation devices of claim 7 to modify analyte, it is included in liquid holding zone or analysis area or modification analyte in both.

33. the method for claim 32, wherein the modification of analyte is reversible.

34. the method for claim 32, wherein the modification of analyte is irreversible.

35. change wettable method in one or more zones in the sample presentation devices of claim 1, it comprises by physical stimulation or chemical stimulation or both modifies the sample presentation devices surface, wherein Qu Yu relative wettable is changed.

36. the method for claim 35, wherein the modification on sample presentation devices surface is reversible.

37. the method for claim 35, wherein the modification on sample presentation devices surface is irreversible.

38. use the sample presentation devices of claim 1 to locate the method for one or more samples, wherein one or more samples move to the one or more zones higher with respect to initial contact point wettable from initial contact point.

39. use the sample presentation devices of claim 10 to locate the method for one or more samples, wherein one or more fluid samples move to the one or more zones higher with respect to initial contact point wettable from initial contact point.