CA2217210A1 - Devices and methods for separating cellular components of blood from liquid portion of blood - Google Patents

Devices and methods for separating cellular components of blood from liquid portion of blood Download PDF

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Publication number
CA2217210A1
CA2217210A1 CA002217210A CA2217210A CA2217210A1 CA 2217210 A1 CA2217210 A1 CA 2217210A1 CA 002217210 A CA002217210 A CA 002217210A CA 2217210 A CA2217210 A CA 2217210A CA 2217210 A1 CA2217210 A1 CA 2217210A1
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Prior art keywords
blood
liquid portion
cellular components
porous
matrix
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Abandoned
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CA002217210A
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French (fr)
Inventor
Wayne H. Schrier
Corey L. Jaseph
Ronald J. Schoengold
Franco Ruggeri
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SmithKline Diagnostics Inc
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Individual
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5002—Partitioning blood components
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/483—Physical analysis of biological material
    • G01N33/487—Physical analysis of biological material of liquid biological material
    • G01N33/49—Blood
    • G01N33/491—Blood by separating the blood components
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
    • G01N33/521—Single-layer analytical elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
    • G01N33/525—Multi-layer analytical elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing
    • G01N33/54386—Analytical elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/195—Assays involving biological materials from specific organisms or of a specific nature from bacteria
    • G01N2333/205—Assays involving biological materials from specific organisms or of a specific nature from bacteria from Campylobacter (G)
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/90—Plate chromatography, e.g. thin layer or paper chromatography
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/805—Test papers
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/81—Packaged device or kit
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/962—Prevention or removal of interfering materials or reactants or other treatment to enhance results, e.g. determining or preventing nonspecific binding
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/97—Test strip or test slide
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S436/00—Chemistry: analytical and immunological testing
    • Y10S436/807—Apparatus included in process claim, e.g. physical support structures
    • Y10S436/808—Automated or kit
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S436/00—Chemistry: analytical and immunological testing
    • Y10S436/807—Apparatus included in process claim, e.g. physical support structures
    • Y10S436/81—Tube, bottle, or dipstick
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T436/00—Chemistry: analytical and immunological testing
    • Y10T436/25—Chemistry: analytical and immunological testing including sample preparation
    • Y10T436/25375—Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T436/00—Chemistry: analytical and immunological testing
    • Y10T436/25—Chemistry: analytical and immunological testing including sample preparation
    • Y10T436/25375—Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]
    • Y10T436/255—Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.] including use of a solid sorbent, semipermeable membrane, or liquid extraction

Abstract

A device for the separation of the liquid portion of blood from the cellular com ponents of blood comprising a pad of porous material (12) permeable to the liquid portion of blood but capable of trapping the cellul ar components of blood; a substrate supporting the pad (16); and means (14), attached to the pad, for facilitating the flow of the liquid por tion of the blood: (i) through interstices around the trapped cellular components of the blood and (ii) from the pad of porous material. The s eparation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad of porous material withou t significant hemolysis. The device can be incorporated into a device for the performance of specific binding assays such as immunoassay s. The pad of porous material can contain an agglutinating agent such as a lectin or an anti-blood cell antibody, or a carbohydrate such as mannitol. Other devices and methods for the separation of the liquid portion of blood from the cellular components of the blood are also d isclosed.

Description

W 096/35952 PCTrUS96106398 DEVICES AND METHODS FOR SEPARATING CELLULAR
COMPONENTS OF BLOOD FROM LIQUID PORTION
OF BLOOD

BACKGROUND OF THE INVENTION
This invention is directed to methods and devices for sepa cellular elements of blood from the liquid portion of blood, particularly in connection with the d~Lellllhl~Lion of characteristics of blood samples.
Among the many analytical ~y~L~llls used for detection and/or S ~leL~ ion of analytes, particularly analytes of biological interest, are chromatographic assay systems.
Such chromatographic systems are frequently used by physicians and medical technicians for rapid in-office ~ gnosi~ and thel~ uLic moniL~ g of a variety of conditions and disorders. They are also increasingly used by patients themselves for at-home monitoring of such conditions and disorders.
Among the most important of such systems are the "thin layer"
system in which a solvent moves across a thin, flat absorbent m.~inm Among the most important of tests that can be pc~lrolllled with such thin layer systems are immnnna~s~ys, which depend on the specific interaction between an antigen or hapten and the corresponding antibody to form antigen-antibody complexes.
The antigen to be det~ct.o-l can itself be an antibody, such as in serological assays for H. pylon-specific antibody. In such cases, the antibody to be detectto-l canalso be bound to a specific antigen. AlL~ dLively, the antigen to be rltotecte-l can be ~letecte-l indirectly by using a labeled second antibody that binds to the first ~ 20 antibody to the analyte to be dett-cte~. These immllnoassays as a means for testing for the presence and/or amount of clinically important molecules have been known for some time. As early as 1956, J.M. Singer reported the use of an immnn~-based latex agghltin~tion test for det~cting a factor associated with CA 022l72l0 l997-l0-02 W 096/35952 - PCTrUS96106398 rh~llm~toid arthritis (Singer et al., Am. J. Med. 22:888-892 (1956)).
Tmmllno~s~ys have been used with chromatographic methods and devices; this combination is known as immnnochromatography Tmmlm~clllulllatographic assays fall into two principal categories:
5 "sandwich" and "c~mpetitive," according to the nature of the antigen-antibody complex to be ~l~tecte(l and the sequence of reactions required to produce that complex.
Examples of sandwich immnnn~c.s~ys ~ ro-llled on test strips are described by U.S. Patent No. 4,168,146 to Grubb et al. and U.S. Patent No.
4,366,241 to Tom et al., both of which are incorporated herein by this l~rt:lellce.
In competitive immnno~c~ys, the disclosing reagent is typically coupled to an analyte or analyte analog which competes for binding with an antibody with any unlabeled analyte present in this sample. Competitive immnnt-assays are typically used for detection of analytes such as haptens, eachlS hapten being monovalent and capable of binding only one antibody molecule.
Examples of haptens include th~ uLic drugs such as theophylline and digoxin and drugs of abuse such as cocaine and heroin and their metabolites. Examples of competitive illllllll.lo~ y devices are those disclosed by U.S. Patent No.
4,235,601 to Deutsch et al., U.S. Patent No. 4,442,204 to Liotta, and U.S.
Patent No. 5,208,535 to Buechler et al., all of which are incorporated herein bythis lcr~ ce.
One of the samples most frequently assayed for an analyte using test strips or similar devices is blood. Most typically, the analyte to be assayed is a soluble component in the liquid portion of blood, i.e., serum or plasma.
The compositions of the two are similar, except that serum, obtained from a blood sample that has been allowed to clot, is lacking in fibrinogen and certainother clotting factors that are depleted as a result of the clotting process.
Most typically, the clinician or technician will draw a blood sample, which is often a fairly small sample. It would be preferable to be able to use the entire blood sample for the assay, avoiding the n~ocPs~ity of a bulk preparation of serum or plasma from the blood sample. However, with most test strips and similar analytical devices, the use of whole blood as a sample, or even W 096/35952 PCTrUS96/06398 3 . .
a blood sample from which the cells, particularly the erythrocytes, have been partially removed, is undesirable.
The blood cells, particularly the erythrocytes, first slow the flow of serum or plasma along the membrane and llltim:~tely stop it by clogging the 5 pores of the membrane. This results in an invalid test. The migration of red blood cells or other blood cells can also create high backgrounds or otherwise h,~lrel~ with the pelrol.llallce of the test carried out by the assay device.
Although blood cells can be removed by filtration through microporous filters, the action of such filters is generally too slow to permit efficient assay of cell-10 free blood.
Additionally, even if the blood cells are effectively removed,methods for doing so frequently result in hemolysis. The occurrence of hemolysis is undesirable because it results in the release of enzymes, hemoglobin, other pigmentc, and stromata into the cell-free portion of blood.
15 This causes hl~elr~lence with many clinical tests.
Various methods for the separation of blood cells from the liquid portion of blood are described, for example in U.S. Patent No. 3,768,978 to Grubb et al., U.S. Patent No. 3,902,964 to Greenspan, U.S. Patent No.
4,477,575 to Vogel et al., U.S. Patent No. 4,594,372 to Zuk, U.S. Patent No.
20 4,753,776 to l:~illm~n et al., U.S. Patent No. 4,816,224 to Vogel et al., U.S.
Patent No. 4,933,092 to Aunet et al., U.S. Patent No. 5,055,195 to Trasch et al., U.S. Patent No. 5,064,541 to Jeng et al., U.S. Patent No. 5,076,925 to Roesink et al., U.S. Patent No. 5,118,428 to Sand et al., U.S. Patent No.
5,118,472 to Tanaka et al., U.S. Patent No. 5,130,258 to Makino et al., U.S.
25 Patent No. 5,135,719 to ~illm~n et al., U.S. Patent No. 5,209,904 to Forney et al., U.S. Patent No. 5,212,060 to Maddox et al., U.S. Patent No. 5,240,862 to Koenhen et al., U.S. Patent No. 5,262,067 to Wilk et al., U.S. Patent No. 5,306,623 to Kiser et al., U.S. Patent No. 5,364,533 to Ogura et al., and U.S. Patent No. 5,397,479 to Kass et al., all of which are incorporated herein by 30 this l~f~l~llce.

W 096/3S952 PCT~US96/06398 However, there is still a need for an improved method of separation of the cellular components of blood from the liquid portion of blood for rapid and accurate assay of analytes contained in the liquid portion of blood.
Particularly, there is a need for an integrated device that incorporates both anS assay element and means for se~ i..g the liquid portions of blood from the cellular components of blood so that an analyte present in the liquid portions of blood can be assayed readily in a single device. Such an improved device would avoid the nlocec~ity of a prelh~ Laly extraction of serum or plasma with its attendant n~ce~ity of safe disposal of the blood fractions. This has become a 10 serious problem due to the increased spread of blood-borne tli~e~es such as hepatitis and AIDS. An improved device would be capable of direct assay of the desired analyte when a whole blood sample is applied to the device.
Preferably, such a device should be able to perform a broad range of immlmo~ ys, including both sandwich and c~ LiLiv~ immnno~ ys.
SUMMARY
We have developed devices and methods for s~ d~hlg the liquid portions of whole blood from the cellular components of blood, as well as assay devices and methods for their use, that meet these needs.
One aspect of the device is a device for st;~dldLillg the liquid portion of blood from the cellular components of blood comprising:
(1) a pad of porous material permeable to the liquid portion of blood but capable of trapping the cellular components of blood;
(2) a substrate supporting the pad; and (3) means, att~r-h~ocl to the pad, for facilit~tin~ the flow of the liquid portion of the blood: (i) through interstices around the trapped cellularcomponents of the blood within the pad and (ii) from the pad of porous material.The separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad without .~ifrnifi~nt hemolysis.

W 096/359S2 PCTrUS96/06398 Typically, the pad of porous material contains a binder for the cellular components of blood. If the binder is an anti-blood cell antibody, it is preferably an anti-erythrocyte antibody. If the binder is a lectin, a number of types of lectins are suitable for use.
O S .~ iv~ly, the pad can be impregn~t~d with a carbohydrate capable of aggregating blood cells. A number of carbohydrates are suitable for use. Preferably, the carbohydrate is m~nnitol.
The pad of porous material in this device can include two sectors:
(i) a first sector permeable to both the liquid portion of blood and the cellular components of blood; and (ii) a second sector permeable to the liquid portion ofblood but capable of binding the cellular components of blood.
Allel.ldlively, the pad of porous material permeable to the liquid portion of blood but capable of trapping the cellular components of blood can include therein an asymmetric membrane with a first surface and a second surface, the membrane having a gradient of pore sizes such that the pore size decreases from the first surface to the second surface, the asymmetric membrane capable of trapping the cellular components of blood within it and allowing the liquid components of blood to pass through.
The means, ~tt~rh~l to the pad, for facilit~ting the flow of the liquid portion of the blood typically includes therein a membrane for cl~ollldtographic separation; the membrane for chromatographic separation typically has a capture zone thereon for binding a member of a specific binding pair.
This device, and other analogous devices described below, can be 25 used in a method of sel)dldlillg the liquid portion of blood from the cellular components of blood. If a membrane for clm~".aLographic s~a,dlion is included, the device can be used in a method for pe,ro,..li..g an assay for detecting and/or d~ ..,i..i..g at least one analyte in the liquid portion of a blood sample.
One aspect of the present invention is a device for s~a.d~ing the liquid portion of blood from the cellular components of blood comprising:

W096/359S2 PCT~US96/06398 (1) a first porous sepaldLing matrix permeable to the liquid portion of blood but capable of Lld~ing the cellular components of blood; and (2) a second porous matrix in operable contact with the first porous sepa~ g matrix that permits the liquid portion of the blood to flow by 5 capillary action or clllullldLographic s~dldLion through the second porous matrix.
The separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the first and second matrices without significant hemolysis.
In this version of the device according to the present invention, the 10 second matrix is typically a membrane for chromatographic st;paldlion, thus producing an assay device. The membrane for chromatographic separation typically has a capture zone thereon for binding a member of a specific binding pair.
If the second matrix is a membrane for chlOlllat~graphic 15 sepdldlion7 a method for p~lrl~llllil.g an assay for ~ietecting and/or ~ . ,t-ii~ at least one dnalyte in the liquid portion of a blood sample can comprise the stepsof:
(1) applying a sample of blood to the first porous s~aldLillg matrix of the device;
(2) allowing the blood sample to flow through the first porous ~epaldlillg matrix to S~hl~l~ the liquid portion of the blood sample from the cellular components of the blood sample;
(3) facilh~ting the flow of the liquid portion of the blood through the interstices around the trapped cellular components of the blood as a result of the action of the second matrix; and (4) allowing the liquid portion of the blood to flow through the second matrix so that an assay is peRormed in the second matrix, the assay beingpeRormed by binding a member of a specific binding pair to the capture zone of the second matrix to detect and/or determine the at least one analyte.
The first sep~r~fing matrix can be an asymmetric membrane with a first surface and a second suRace. The membrane has a gradient of pore sizes such that the pore size decreases from the first suRace to the second suRace; the a~ylllllletric membrane is capable of trapping the cellular components of blood within it and allowing the liquid components of blood to pass through.
Typically, the device further comprises an impermeable solid support to which the second matrix is fixedly ~tt,l~h~od Yet another aspect of the invention is a device for the separation of the liquid portion of blood from the cellular components of blood CUlll~liSi~g three matrices. Such a device can clJlll~lise:
(1) a first porous s~aldLhlg matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood;
(2) a second porous ~epaldLillg matrix in operable contact with the first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (3) a third porous matrix in operable contact with the second porous sepalalillg matrix that permits the liquid portion of the blood to flow by 15 capillary action or chlollla~graphic separation through the second porous matrix.
The separation of the liquid portion of the blood from the cellular components of the blood occurs by flow through the first and second porous s~dldLillg matrices without !sipnifir,Jnt hemolysis.
Yet another embodiment of a device according to the present 20 invention has mllltirle second porous matrices. Such a device comprises:
(1) a first porous s~aldLillg matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (2) at least two second porous m,1trirçs, each second porous matrix in operable contact with the first porous st~aldLillg matrix that permits the 25 liquid portion of the blood to flow by capillary action or chromatographic separation through the second porous matrix.
Still another aspect of the present invention is a two-component device for s~aldlillg the liquid portion of a blood sample from the cellular components. This device comprises:
(1) a first opposable component including:
(a) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and W O 96/35952 PCTrUS96/06398 (b) a second porous matrix in operable contact with the first porous sepalalillg matrix that permits the liquid portion of the blood to flow by capillary action or chromatographic separation through the second porous matrix;and (2) a second opposable component att~rh~hle to the first opposable component such that the first and second opposable components can be brought into opposition to transfer fluid from one of the opposable components to the other by pressure.
The separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the first and second matrices of the first opposable component without ~ignifir~nt hemolysis.
The second opposable component can include a sample preparation zone, which can include at least one reagent for tre~tmPnt of the sample or a specific binding partner labeled with a ~iPtPct~hle label, the specific binding partner having specific binding affinity for at least one component selected from the analyte and a specific binding partner for the analyte in a form that can beresolubilized by the addition of an aqueous sample to the sample pL~dldLion zone.
A two-component device particularly adapted to bidirectional assays can comprise:
(1) a first opposable component including:
(a) a first porous s~,a~li..g matrix permeable to the liquid portion of blood but capable of LLd~pillg the cellular components of blood; and (b) a second porous matrix inrl~lrling a membrane for 25 chromatographic separation in operable contact with the first porous separating matrix that permits the liquid portion of the blood to flow in a first direction by capillary action or chromatographic sepaldLion through the second porous matrix;and (2) a second opposable component ~tt~rll~hle to the first opposable 30 component such that the first and second opposable components can be brought into opposition to transfer a reagent from the second opposable component to thefirst opposable component by pressure such that bringing the first and second W 096/35952 PCT~US96/06398 opposable components into opposition causes the reagent transferred from the second opposable component to the first opposable component to migrate through the second porous matrix in a second direction opposite to the first direction.
In this version, the s~aldLion of the liquid portion of blood from S the cellular components of the blood occurs by flow through the first and second matrices of the first opposable component without ~i~nific~nt hemolysis.
Another aspect of the present invention is a method for separating the liquid portion of blood from the cellular components of blood comprising thesteps of (1) adding a cross-linking substance for the cellular components of blood to a sample of whole blood, the cross-linking substance being selected from the group con.ci.cting of a lectin, an anti-blood cell antibody, and a carbohydrate capable of ag~.~galillg blood cells;
(2) mixing the cross-linking substance and the blood sample to 15 form a ~ Lulc of the cross-linking sl~bst~nre and the blood sample;
(3) applying the llli~lUlC of the cross-linking substance and the blood sample to a device for S~dld~ g the liquid portion of blood from the cellular components of blood, the device colll~lising.
(a) a pad of porous m~t-o,ri~l permeable to the liquid portion 20 of blood but capable of trapping the cellular components of blood aggregated by the reaction between the cross-linking substance and the blood sample;
(b) a substrate supporting the pad; and (c) means, ~tt~rh~1 to the pad, for facilit~ting the flow of the liquid portion of the blood: (i) through interstices around the trapped cellular 25 components of the blood and (ii) from the pad of porous material, whereby theseparation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad without ~ignifir.~nt hemolysis; and (d) allowing the liquid portion of the blood to flow through the pad to separate the liquid portion of the blood from the cellular components of 30 the blood.

W096/35952 PCTrUS96/06398 Preferably, the method further comprises adding an anticoagulant together with the cross-linking substance. Typically, the anticoagulant is heparin or EDTA.
Preferably, a concentration of cross-linking substance is used that 5 is sufficient to cross-link subst~nfi~lly all cellular elem~qnt~ of blood.
An alL~lllaLive method for sepal~ g the liquid portion of blood from the cellular components of blood comprises the steps of:
(1) adding a sample of blood to a capillary tube coated with a cross-linking s-7bst,7nl~e as described above;
(2) allowing and cross-linking substance to dissolve in the blood sample to form a mixture of the cross-linking substance and the blood sample;
(3) applying the llli~LUl~ of the cross-linking substance and the blood sample to a device for separating the liquid portion of blood from the cellular components of blood as described above; and (4) allowing the liquid portion of the blood to flow through the pad to separate the liquid portion of the blood from the cellular components of the blood.
Preferably, the capillary tube is also coated with an anti-coagulant.
BRIEF DESCRIPTION OF THE DRAWINGS
These and other features, aspects, and advantages of the present invention will become better understood with reference to the following description, appended claims, and accolll~llying drawings where:
Figure 1 is a drawing of a device for se~al~Lillg t7.1e liquid portion of blood from the cellular components of blood, employing a pad of porous material;
Figure 2 is another drawing of the device shown in Figure 1, showing the migration of blood through the device;
Figure 3 is a drawing of another embodiment of an assay device according to the present invention employing a porous pad with two sectors;
--W 096/35952 PCTrUS96/06398 Figure 4 is a drawing of another embodiment of a device for ~t;pa.clLillg the liquid portion of blood from the cellular components of blood,employing three matrices;
Figure 5 is a drawing of yet another embodiment of an assay device according to the present invention with two second matrices, which can incorporate assay elements;
Figure 6 is a drawing of an embodiment of a two-component device according to the present invention;
Figure 7 is a drawing of another embodiment of a two-component device according to the present invention; and Figure 8 is a scllem~tic diagram of a method according to the present invention for s~alaLillg the liquid portion of blood from the cellular components of blood, employing off-board separation, with blood being added to a capillary tube co.ll;.i.~ g a cross-linking substance for the cellular components of blood.

DESCRIPTION

Definitions In the context of this disclosure, the following terms are defined as follows unless otherwise intlil~t~cl Specific Binding Partner: A member of a pair of molecules that interact by means of specific non-covalent interactions that depend on the three-tlimPn~ional structures of the molecules involved. Typical pairs of specific binding l,~.Ll~ include antigen-antibody, hapten-antibody, hormone-receptor, nucleic acid strand- complem~nt~ry nucleic acid strand, substrate-enzyme, substrate analog-enzyme, inhibitor-enzyme, carbohydrate-lectin, biotin-avidin, ~ and virus-cellular receptor.
Operable Contact: Two solid components are in operable contact when they are in contact, either directly or indirectly, in such a manner that an aqueous liquid can flow from one of the two components to the other subst~nti~lly ~ . . uyL~dly, by capillarity or otherwise. "Direct contact" means W 096/35952 PCTrUS96/06398 that the two elements are in physical contact, such as edge-to-edge or front-to-back. Typically, when two components are in direct contact, they are overlapped with an overlap of about 0.5 mm to about 5 mm. However, the components can be placed with abutting edges. "Indirect contact" means that the two elemen 5 are not in physical contact, but are bridged by one or more conductors.
Analyte: The term "analyte" includes both the actual molecule to be assayed and analogs and derivatives thereof when such analogs and derivativesbind another molecule used in the assay in a nla~ subst~nti~lly equivalent to that of the analyte itself.
Antibody: The term "antibody" includes both intact antibody molecules of the a~ v~lial~ specificity and antibody fr~gments (including Fab, F(ab'), and F(ab')2 fr~gmentc) as well as ch.?mi~ ~lly modified intact antibody molecules and antibody fr~gmto-nts, including hybrid antibodies assembled by in vitro reassociation of subunits and single-chain antibody molecules produced by 15 genetic ergin~ering. Also included within the definition are anti-idiotypic antibodies that specifically bind antigen-combining sites of antibodies.
Without Signifir~nt Hemolysis: The term "without .signific~nt hemolysis" means the absence of hemolysis to a degree such that the rçs-llting plasma or serum shows no ~ al~llt redness against a white background by visual 20 inspection.
Supported: The term "~uppolLed" can include either directly or indirectly supported, as directly by a solid substrate or indirectly by a solid substrate through one or more ill~ /ellillg elem~nt~.
Cross-linking Substance: The term "cross-linking substance" is 25 used generically herein to include substances that are capable of cross-linking, ag~ l ;, .g, or aggregating the cellular components of blood. Specifically, thisterm includes lectins and anti-blood cell antibodies, as well as carbohydrates that can aggregate blood cells by making them adhesive and causing them to clump.
Methods and devices according to the present invention use one of 30 two tech~iques for separating the cellular elements (formed elements) of blood from the liquid portion of blood (serum or plasma, co..~ i"g the soluble elements), for use in an immllnochromatographic test format.

CA 022l72l0 l997-l0-02 W 096/3S952 PCTrUS96/06398 The first of these techniques is active separation of the cellular elements of blood from the liquid portion of blood on or as an integral part of the test device, referred to generally as on-board procee.einp~. The second of these techniqlles is sepal~lion or blood sample proce.eeing before the sample is5 added to the test device, referred to generally as off-board proce.c.eing.

I. DEVICES AND METHODS FOR ON-BOARD PROCESSING
A. General Description of On-Board Processin~
One aspect of the present invention is devices for St~ Lldlillg the 10 liquid portion of blood from the cellular components of blood on or as an integral part of the test device. The cellular components of blood include erythrocytes (red blood cells), leukocytes (white blood cells) and platelets. The liquid portion of the blood includes the r~Tn~in~ltor of the blood and is generally known as serum if the blood has clotted, follllhlg a clot co..~ .i..g fibrin and the 15 blood cells. It is generally known as plasma if it is obtained from unclottedblood. The major ingredient present in plasma but absent in serum is fibrinogen,the precursor of fibrin.
In general, such a device comprises:
(1) a pad of porous material permeable to the liquid portion of 20 blood but capable of trapping the cellular components of blood;
(2) a substrate supporting the pad; and (3) means, ~tt~r.h~l to the pad, for facilit~ting the flow of the liquid portion of the blood: (i) through interstices around the trapped cellularcomponents of the blood and (ii) from the pad of porous material.
In general, a method for s~L)al~lillg the liquid portion of blood from the cellular components of blood using this device comprises:
(a) applying a sample of blood to the pad of porous material of - the device;
(b) allowing the blood sample to flow through the pad of porous 30 material to separate the liquid portion of the blood sample from the cellular components of the blood sample; and CA 022l72l0 l997-l0-02 W 096/35952 PCTrUS96/06398 (c) facilit~ting the flow of the liquid portion of the blood through the interstices around the trapped cellular colllpollellL~ of the blood and from the pad of porous material.
Various a~ gements and elaborations of this device are within 5 the scope of the present invention, as described further below.
The separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad without ~ignific~nt hemolysis.
Typically, the substrate is a solid, substantially planar substrate.
10 Typically, the flow through the pad occurs in a direction subst~nti~lly parallel to or along the substrate.
The means, ~tt~rhP(l to the pad, for facilit~ting the flow of the liquid portion of the blood can include therein a membrane for chromatographic sepal~Lion; typically, the membrane has a capture zone thereon for binding a 15 member of a specific binding pair. In this arrangement, the device can be used in a method for pelrolllli~lg an assay for ~letPcting and/or de~t- .. .i..i . .g at least one analyte in the liquid portion of a blood sample colllL.li~ g the steps of:
(1) applying a sample of blood to the pad of porous material of the device;
(2) allowing the blood sample to flow through the pad of porous material to separate the liquid portion of the blood sample from the cellular components of the blood sample;
(3) facilit~ting the flow of the liquid portion of the blood through the interstices around the trapped cellular components of the blood as a result of the action of the means attached to the pad; and (4) allowing the liquid portion of the blood to flow through the chromatographic mP~linm so that an assay is performed in the chromatographic medium, the assay being ~lrol.lled by binding a member of a specific binding pair to the capture zone of the chl~ graphic m~ m to detect and/or ~leterminP the at least one analyte.

W O 96/35952 PCT~US96/06398 The conditions that are optimal for the performance of such assays, such as the choice of the member of the specific binding pair, the use of buffers or salts, the time required, and the oplilllulll temperature, are well known in the art and need not be described further here.
The porous pad, also referred to as a sample pad because the sample is typically applied to it, can be a woven or non-woven fabric, paper, cellulose, glass fiber, polyester, other polymers, or llli~LulcS of these materials to retain the cellular components of blood. The porous pad typically has a binder for the cellular components of blood incorporated therein.
The binder for the cellular components of blood is typically a lectin or an anti-blood cell antibody. When the binder is an anti-blood cell antibody, it is typically an anti-erythrocyte antibody. Such antibodies are wellknown in the art and need not be described further here. Typically, they are obtained by the injection of red blood cells or fractions from red blood cells into a dirr~l~llL species. If the desired antibody is anti-human red blood cell antibody, suitable ~nim~l~ for the production of such antibodies include goats, rabbits, horses, and sheep. Either polyclonal or monoclonal antibodies can be used.
AlLc~ Liv~ly, anti-leukocyte or anti-platelet antibodies can be used alone or inaddition to the anti-red blood cell antibody if it is desired to ensure removal of those cellular components.
The binder for the cellular components of blood can be noncovalently bound to the sample pad. AlLelll~Livcly, it can be covalently cross-linked to the sample pad; techniques for cross-linking pl~teills to solid supports such as cellulose, paper, and other typical sample pad materials are well-known in the art and need not be described further here. The sample pad, cont~ining antibodies or lectins, can be further treated with polyester binders to capture cellular elements, as described, for example, in U.S. Patent No. 4,816,224 to ~ Vogel et al., incorporated herein by lcrclcnce. Other types of polymer binders can also be used.
When the binder is a lectin, typically the lectin is one of the following, but is not limited to: concanavalin A, abrin, phytoh~ ggluLu limlllin, or one of the lectins produced by the following species: Agaricus W 096/35952 PCTrUS96/06398 bisporus, Anguilla anguilla, Arachis hypogaea, Bandeiraea simplicifolia, Bauhinia purpurea, Caragana arborescens, Cicer arietinum, Codium fragile, Datura stramonium, Dolichos biflorus, Erythrina corallodendron, Erythrina cristagalli, Euonymus europaeus, Glycine max, Helix aspersa, Helix pomatia, 5 Lathyrus odoratus, Lens culinaris, Tycopersicon esculentum, Maclura pomifera, Momordica charantia, Mycoplasma gallisepticum, Naja mocambique, Naja kaouthia, Perseau americana, Phaseolus coccineus, Phaseolus limensis, Phaseolus vulgaris, Phytolacca americana, Pisum sativum, Pseudomonas aeruginosa, Psophocarpus tetragonolobus, Ptilota plumosa, Ricinus communis, Robinia pseudoacacia, Sambucus nigra, Solanum tuberosum, Sophora japonica, Tetragonolobus purpureas, Triticum vulgaris, Ulex europaeus, Vicia faba, Vicia sativa, Vicia villosa, Vigna radiata, Viscum album, and Wisteria floribunda.
Lectins are l.rot~ ls produced by plants and some animal species that bind specifically and non-covalently to sugar groups that are present on the surface of 15 blood cells.
Preferably, the lectin is capable of binding both erythrocytes and leukocytes and is not blood-cell-group specific. Many other examples of lectins are known and need not be described further here.
The pad of porous material can ~ltçrn~ively be impregnated with a 20 carbohydrate capable of aggregating blood cells, such as the carbohydldl~s disclosed in U.S. Patent No. 4,678,757 by Rapkin et al., incorporated herein by this lcrelcnce. These carbohydrates include, but are not npcç~s~rily limited to,ikl, sorbitol, inositol"B-D-glucose, c~-D-glucose, D(+)xylose, D(+)m~nn~-se, D(-)arabinose, L(+)arabinose, D(+)galactose, L(-)xylose, D-25 glucoheptose, L-lyxose, lactose, maltose, and sucrose. A particularly ~r~r~ dcarbohydrate is ..~ h Although applicants do not intend to be bound by this theory, these carbohydrates are believed to act by binding non-covalently to thesurface of erythrocytes, making them adhesive and causing them to clump or aggregate.
A carbohydrate in solution is applied to a permeable matrix such as a non-woven fiber (e.g., cellulose, glass, or polyester) in a concentration up to 20% (w/v) to produce a treated matrix. The solution can be applied by various means such as impregnation, printing, or spraying to achieve the desired concel~Lldlion in the matrix. The carbohydrate functions as a holding, clumping,or aggl~ g agent which 3~3lef~l~llLially separates cells from the ~3u~ u,ldmg liquid which is free to migrate through the matrix.
3 5 The volume of blood separated is a function of the absorbentcapacity of the treated matrix, the means, attz3~hen to the pad, for facilitzting the flow of the liquid portion of the blood through interstices around the trapped cellular components of the blood and from the pad, and the degree and area of adherence between the treated matrix and the means for facilitztin~ the flow of the liquid portion of blood.

B. Particular Embon3imentc of Devices for On-Board Processin~
One embodiment of a device for on-board procec.cing according to the present invention comprises:
(1) a first porous s~3~3dldLillg matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (2) a second porous matrix in operable contact with the first porous s~dldlillg matrix that permits the liquid portion of the blood to flow bycapillary action or chromatographic separation through the second porous matrix without cignifirzlnt hemolysis.
In this embodiment, the second porous matrix comprises the means, zttz3.eh~0(i to the pad, for facilitz3.ting the flow of the liquid portion of the blood through interstices around the trapped cellular components of the blood and from the pad of porous material. The second porous matrix can be a membrane such as a membrane suitable for chromatographic s~3~3aldLion. Typical materials for such membranes include, but are not limited to, nitrocellulose, cellulose, other cellulose derivatives, nylon, rayon, paper, silica, polyesters, and polysulfones. A generally 3~3lert;ll~d material for such membranes is nitrocellulose. The cL.~,lllaLographic mf~-~.3illm can be pleLlGat~d or modified as neer~en3 CA 022l72l0 l997-l0-02 W096/35952 PCTrUS96/06398 This second porous matrix can have capture zones thereon for binding members of a specific binding pair, such as antigens, haptens, or antibodies. For example, the second porous matrix can have, immobilized in the capture zone, a first antibody for binding analyte, which is then detected by means of a labeled second antibody in a sandwich reaction. ~ liv~ly, the second porous matrix can have an antigen immobilized in the capture zone for binding of an antibody. More than one capture zone can be present on the same second porous matrix; if more than one capture zone is present, they can have the same or different members of a specific binding pair bound thereto. If more 10 than one capture zone is present, one capture zone can be used as a control to ensure that the assay has been performed properly. Many arrangements are well known in the art and need not be recited further. The second porous matrix can therefore comprise a chromatographic assay element, which can be used for the performance of an immlln~chromatographic assay. When the second porous 15 matrix is a chromatographic assay element, the device is capable of pelrollnillg on-board separation of the cellular components of the blood from the liquid portion of the blood and an assay for an analyte in the liquid portion of the blood in a unitary device. The assay can be carried out by applying the blood sample to the first s~al~Lillg matrix and subsequently reading the result.
Typically, the chromatographic assay element pelrolll-s either a competitive immlm~assay or a sandwich i"""~",o~ y, as these formats are generally known in the art.
The labeled component bound to the chromatographic m~-linm, in the case of a sandwich immlmoassay, is typically a labeled antibody to the 25 analyte. If the analyte is itself an antibody, such as in the case of an assay for the ~lett-cti-)n of antibody in human serum to the bacterium Helicobacter pylori, suspected of being the causative agent of stomach ulcers, the labeled component can be a second antibody that binds the first antibody on the basis of species, class, or subclass specificity. Class specificity is also known as isotype 30 specificity, such as IgG, IgM, IgA, IgD, and IgE for human antibodies.
Subclass specificity refers to antigenic differences within classes, such as IgGl, IgG2, IgG3, and IgG4, which are subclasses of IgG. It is highly plc~ d that W 096/359S2 PCTrUS96106398 the labeled specific binding partner used for detection of an antibody analyte binds to the constant region of the antibody analyte, in order to prevent illL~lr~lcllCe.
In some applications, it is desirable to employ indirect labeling.
5 For example, in testing for Giardia antigen, an IgM antibody can be used that may be difficult to label directly. In that case, a secondary specific binding partner specific for the mobile first specific binding partner can be labeled.
Typically, the labeled secondary specific binding partner binds to the antibody that is the first specific binding partner on the basis of species, class, or subclass 10 specificity. The first specific binding partner has specific binding affinity for the analyte. As an alternative to the use of a secondary specific binding partner, the first specific binding partner can be conjugated to biotin and an avidin-conjugated label can be used.
When a competitive immnnn~say is performed, the label is 15 typically an analyte or analyte analog. However, other labeling schemes are known in the art; in some of these labeling sch~mes7 the label is a labeled antibody to the analyte or a secondary specific binding partner. In some cases, anti-idiotypic antibodies can be used for colll~LiLiv~im mlm~assays.
An additional elem~nt or eleTn~nt.c can be interposed between the 20 first porous s~aldLhlg matrix and the second porous matrix. These element.~, which are typically con~ rting~ can act as a bridge between the first porous s~dl~Lillg matrix and the second porous matrix, i.e, the chromatographic assay element.
Optionally, and preferably, the second porous matrix is fixedly 25 ~tt~rhr~l to a solid support that is impermrahle. The second porous matrix can be l~min~t-orl to the support or cast onto it. The solid support can be made of materials such as plastic or l~ r(l cardboard.
Such a device is shown in Figure 1. The device 10 includes a first porous s~l!ala~ g matrix 12, a second porous matrix 14 in operable contact with 30 the first porous sepdldlillg matrix 12, and a solid support 16. The first porous separating matrix 12 has a first surface 18 and a second surface 20. The second porous matrix 14 can be a chromatographic assay element.

W 096/35952 PCTrUS96/06398 In use, a blood sample 22 is added to the first surface 18 of the first porous st:~dldLillg matrix 12, and the liquid portion of the blood sample 22 migrates into the second porous matrix 14 as a result of the contact between thesecond surface 20 of the first porous sel)al~ g matrix 12 and the second porous 5 matrix 14 after the cellular elements are trapped within the first porous separating matrix 12. A chromatographic assay can be performed within the second porous matrix 14.
Figure 2 shows the device of Figure 1 after the liquid portion of the blood sample has migrated into the second porous matrix 14. The cross-10 h~t~h~-l regions in Figure 2 represent the areas of liquid flow through the first porous matrix 12 and the second porous matrix 14.
In an ~lle~;-liv~ version of this embodiment, the first separating matrix can be an ullL~ d asymmetric membrane. The untreated asymmetric membrane is constructed in such a way that it has a decreasing gradient of pore 15 size within the membrane. The asymm--tric membrane has a first surface and a second surface; the blood sample is applied to the first surface. The pore size decreases from the first surface to the second surface. The asymm~tric membrane is capable of trapping the cellular components of blood within it and allowing the liquid components of blood to pass through. The first ~al~lhlg 20 matrix permits the liquid portion of the blood to flow through on contact with the second matrix, as described above.
This device is also depicted by the drawings of Figures 1 and 2, with the first surface 18 and second surface 20 of the asymm~tric membrane as the first porous separation matrix 12. Blood flow is from the first surface 18 to 25 the second surface 20 of the asymmetric membrane.
Asymmetric membranes suitable for use in on-board s~palaLillg devices according to the present invention can be prepared from combin~tion~ of hydrophobic and hydrophilic polymers, such as disclosed in U.S. Patent No.
5,240,862 to Koenhen et al. and U.S. Patent No. 5,076,925 to Roesink et al.
30 The hydrophobic polymer can be polysulfone, polyether sulfone, polyimide, or polyetherimide, and the hydrophilic polymer can be polyvinyl pyrrolidone, polyacrylic acid, polyvinyl alcohol, polyvinyl acetate, or polyethylene glycol.

CA 022l72l0 l997-l0-02 W 096/35952 PCTrUS96/06398 In yet another alLclllaLivc version of this embo-lim~nt the first matrix can be constructed in such a way that only a portion of the pad is capable of binding the cellular components of blood. In other words, the pad can be divided into two sectors, a first sector that permits flow but is not capable ofbinding the cellular components of blood and a second sector that is capable of bindillg the cellular components of blood. The second sector can contain antibodies, lectins, or carbohydrates as described above. The first sector typically contains reagents for ~lcll~ -ent of the blood sample that can be premixed into the blood sample when the blood migrates through the first sector.This alLcllla~ive version of the device is depicted in Figure 3. The device 40 has a first sepaldLillg matrix 42 with a first surface 44 and a secondsurface 46, with two sectors, a first sector 48 that is not capable of binding the cellular components of blood and a second sector 50 that is capable of binding the cellular components of blood. The device also has a second porous matrix 52 and a solid support 54.
In use, a blood sample 56 iS added to the first surface 44 of the first scpala~illg matrix 42, and it migrates from the first sector 48 to the second sector 50, typically premixing the reagents present in the first sector 48 into the blood sample 56 for pretre~tn ent of the blood sample 56. The liquid portion of the blood sample 56 then migrates to the second porous matrix 52 from the second sector 50; a clnulllatographic assay can be l?elrclmed in the second porous matrix 52.
In still another alL~llative version of this embo-liment, three elements are used:
(1) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood;
(2) a second porous separating matrix in operable contact with the first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (3) a third porous matrix in operable contact with the second porous ~cl!ald~ g matrix that permits the liquid portion of the blood to flow bycapillary action or chromatographic sepdldLion through the second porous matrix.

W096/35952 PCTrUS96/06398 The separation of the liquid portion of the blood from the cellular components of the blood occurs by flow through the first and second porous se~aldLhlg matrices without ~i~nific~nt hemolysis.
In this ~lttorn~tive version, the third matrix comprises the means 5 for facilit~ting the flow of the liquid portion of the blood: (i) through interstices around the trapped cellular components of the blood and (ii) from the second matrix. The third matrix can include a chromatographic assay element.
Optionally, and preferably, the third matrix is fixedly ~tt~ch~ to a solid support that is impermeable, as described above.
The first and second matrices can be alike or different; they can co,.lplise any of the alL~ dLives described above in Section I(B), including matrices cont~ining a binder for the cellular c~nll-ollellL~, of blood such as a lectin or an anti-blood cell antibody, matrices cont~ining a carbohydrate capable of aggregating blood cells, and matrices cont~ining an asymmetric membrane to trap blood cells. Matrices with two sectors can be used.
For this alLt:lllaLive of the device, a method for s~aldLillg the liquid portion of blood from the cellular components of blood can comprise the steps of:
(1) applying a sample of blood to the first porous s~aldLhlg matrix of the device;
(2) allowing the blood sample to flow through the first porous sepaldLillg matrix and the second porous s~aldLillg matrix to separate the liquid portion of the blood sample from the cellular components of the blood sample;
and (3) facilh~ting the flow of the liquid portion of the blood through the interstices around the trapped cellular components of the blood as a result of the action of the third matrix.
When the third matrix includes a membrane for chromatographic separation with a capture zone, a method for performing an assay for (let~cting 30 and/or detennining at least one analyte in the liquid portion of a blood sample can comprise the steps of:

W 096/3S952 PCT~US96/06398 (1) applying a sample of blood to the first porous separating matrix of the device;
(2) allowing the blood sample to flow through the first and second porous s~dldlillg matrices to separate the liquid portion of the blood sample 5 from the cellular components of the blood sample;
(3) facilit~fin~ the flow of the liquid portion of the blood through the interstices around the trapped cellular components of the blood as a result of the action of the third matrix and;
(4) allowing the liquid portion of the blood to flow through the 10 third matrix so that an assay is performed in the second matrix, the assay being performed by binding a member of a specific binding pair to the capture zone of the third matrix to detect and/or del~lllli/le the at least one analyte.
This all~llldlive version of the device is shown in Figure 4. The device 60 has a first matrix 62, a second matrix 64, a third matrix 66, and a 15 solid support 68. A blood sample 70, applied to the first matrix 62, flows through the first matrix 62 and the second matrix 64; the liquid portion of the blood sample then migrates into the third matrix 66. A chromatographic assay can be performed in the third matrix 66.
In yet another alLelllaLivt;: version of this embodiment, the device 20 can include multiple second porous m~trirçs, each second porous matrix being in operable contact with the first porous s~alalillg matrix. Each second porous matrix can comprise a chromatographic assay element, with a capture zone, as described above. When the second porous matrices include chromatographic assay elements with capture zones, a method for performing an assay for 25 detçcting and/or d~l~l Illillil~g at least one analyte in the liquid portion of a blood sample can comprise the steps of:
(1) applying a sample of blood to the first porous s~aldling matrix of the device;
(2) allowing the blood sample to flow through the first porous s~aldLillg matrix to separate the liquid portion of the blood sample from the cellular components of the blood sample;

(3) facilit~tin~ the flow of the liquid portion of the blood through the interstices around the trapped cellular components of the blood as a result of the action of the second matrices; and (4) allowing the liquid portion of the blood to flow through the 5 second matrices so that an assay is performed in at least one of the second matrices, the assay being ~lrolllled by binding a member of a specific binding pair to the capture zone of at least one of the second matrices to detect and/ordetermine the at least one analyte.
In one arrangement, depicted in Figure 5, the device includes two 10 second porous matrices, one in operable contact with each end of the first porous matrix. In this arrangement, the blood is applied near the center of the first porous matrix, and migr~tes uuLw~d to the ends. Alternatively, three or more second porous matrices can be used, each in operable contact with the first porous matrix. The second porous m~trire~ can be arranged ch-;ulllfelellLially 15 around the first porous matrix, like the spokes of a wheel. In this ~ ...,.1iv~, the first porous matrix can be any of the first porous matrices described above,including the untreated asymm~tric membrane.
In Figure 5, the device 80 comprises a first porous separation matrix 82, with a first surface 84 and a second surface 86 and first and second ends 88 and 90, and two second matrices 92 and 94, as well as a solid support 96. The two second matrices 92 and 94 are in contact with the ends 88 and 90 of the first porous separation matrix 82. A blood sample 98 iS added to the first surface 84 of the first porous sepalaLion matrix 82 and migrates through the first porous sepaldLion matrix 82, with the liquid portions of the blood sample migrating into the two second matrices 92 and 94.

C. Two-Component Assay Devices Another embodiment of the present invention is a two-component device incorporating the first and second matrices. Such a device, in general, 30 comprises:
(1) a first opposable component including:
(a) a first porous se~aldLillg matrix as described above; and , porous separaring matrix as descnbed above; and (2) a second opposable component ~ rll~bie to the first opposable component such that the first and second oppossble components can be brou~ht into opposition to transfer fluid firom one of the opposable comr.~n~ntc to ~he o~r by pressure.
A large number of embodim~rltc ex~st using two opposa~e componen~s.
Set~eral ~Itematives are dep~cted below in Figures 6 and 7. These alternatives are eY~rnr]~ry and not exclusive; a large number of fo~ns of the assay device exist, and have 0 been described, for exarnple in cop~r ~in~ U.S Patent ~pplic~tion ~er~al l~o Og~o40,431) ~ncc,~ulat~d herein by this ~efL,~e.,ce.
For exarnple, ~he second opposable co~ F..t can include a samplc application zone, which can then include at least one reagent for contact with the sample.
This reagent can be used for contac~ with the sample bcfore the s~ ion of the liqu~d --~ 15 portio~ of the ~~ood from the cellular cr mpo~ts of the blood.
Altematively, the sample applic~tion zone c3n include a specific bindin~
partne~ labeled with a r~elpc~ble label. The c~~çific bin~ P partner can have specific binding affinity for at least one co~pon~ t select~t ~om the analyte and a sperific bind~ng partner for the a~alyte in a fonn that can be resolubilized by the addition of an 2 0 aqueous sarnple to the sample ~lc~ ion zone. In other words, the labeled specific binding partner can be applied to the sample application zone in liquid form and dried down ~n such a wsy Ihs~ be redissolved. Typically, whe:n the device is used for a sandw~ch imrn~ s~y~ the specific binrlinf~ partner labeled with the detectable label has ~r~ific bin~ aff~ty for the analyte, -- 2 5 Alte~ ivcly, the first opposable cG~pol~ent can fi~rther inclute a s rnple application zone, which can include a spe~ifir binding partner labeled with a deeectable label in resolubilizable fom~. As indicated below, ~ this cas~, the ~mplc applicali~n zone on the first opposable component would be contac~ed by an elemen~ on the second opposable comrone-lt when the first and second opposable cG~ o~ents are brought in~o 3 0 oppo~ilion. 1-hi5 will result in the q CA 02217210 i997-10-02 W 096/3S952 PCTrUS96/06398 Lldn~rel of the sample; the sample and the resolubilized labeled specific binding partner are then applied to the porous pad.
AlL~ a~iv~ly, in a two-component device, the porous pad can be on the opposite component from the chromatographic m~ m An example of this arrangement is shown below in Figure 6.
A method for pelrc,.lllhlg an assay for ~etectin~ and/or determining at least one analyte in the liquid portion of a blood sample can comprise the steps of:
(1) applying a sample of blood to the first porous s~L~alalhlg 10 matrix on the first opposable component of the two-component assay device;
(2) allowing the blood sample to flow through the first porous se~dldLillg matrix to ~epalal~ the liquid portion of the blood sample from the cellular components of the blood sample;
(3) facilit~ting the flow of the liquid portion of the blood through 15 the interstices around the trapped cellular components of the blood as a result of the action of the second matrix;
(4) bringing the first and second opposable components into opposition to Ll~l~rel fluid from one of the opposable components to the other by ~l~S~ul~; and (5) allowing the liquid portion of the blood to flow through the second matrix so that an assay is performed in the second matrix, the assay being performed by binding a member of a specific binding pair to the capture zone of the second matrix to detect and/or determine the at least one analyte.
Several examples of two-component assay devices are shown.
One general arrangement is shown in Figure 6. The assay device 200 has a first opposable component 202 and a second opposable component 204.
The first opposable component 202 includes a porous pad 206 for application of the sample. The second opposable component 204 contains a chromatographic medium 208. The means for withdrawing a liquid portion of blood from the 30 porous material is formed by the overlap between the porous pad 206 and the chromatographic m~ lm 208 when the first and second opposable components 202 and 204 are brought into opposition. The chromatographic m~tlillm 208 can CA 022l72l0 l997-l0-02 W 096/35952 PCTrUS96/06398 include a detection zone 210 and a control zone 212. The first opposable component 202 and the second opposable component 204 are joined by a hinge 214. The chromatographic mP~ lm 208 is supported in a well 216. The first opposable component 202 can include a window 218 for viewing of the S chromatographic m~illm 208, including the area of the detection zone 210 and the control zone 212. The first and second opposable components 202 and 204 can be held together by engagers, such as those formed by a beveled edge 220 on the first opposable component 202 and an undercut edge 222 on the second opposable component 204. Other types of engagers can also be used. The 10 device can be ~ccessed through a notch 224 formed in the second opposable component 204.
Another embodiment of an assay device according to the present invention comprises a device capable of carrying out bidirectional chlo~ ography. This embodiment is shown in Figure 7. The assay device 300 15 has a first opposable component 302 and a second opposable component 304.
The first opposable component 302 includes an absorber 306, which can be an absorbent pad, and an applicator 308. The second opposable component 304 has a chromatographic medium 310 having a first end 312 and a second end 314, with a detection zone 316 and a control zone 318. The second opposable component 304 also has a conductor 320 in operable contact with the second end 314 of the chromatographic medium 310; the conductor 320 is used for application of a reagent in the applicator 308 to the chromatographic medium 310when the first and second opposable components 302 and 304 are brought into opposition. The second opposable component 304 also has a pad of porous material 322 permeable to the liquid portion of blood but capable of binding thecellular components of blood as described above. The pad of porous material 322 iS in operable contact with the first end 312 of the chromatographic mP-lillm 310; this operable contact forms the means for withdrawing the liquid portion ofblood from the pad of porous material 322. The first and second opposable components 302 and 304 are joined by a hinge 324. The chromatographic medium 310 and pad of porous material 322 are supported in a well 326. The first opposable component 302 can include a window 328 for viewing of the CA 022l72l0 l997-l0-02 W 096/3S952 PCTrUS96106398 chromatographic m.oflillm 310, including the area of the detection zone 316 and the control zone 318. The first and second opposable components 302 and 304 can be held together by engagers, such as those formed by a beveled edge 330 on the first opposable component 302 and an undercut edge 332 on the second 5 opposable colllpollellL 304. Other types of engagers can also be used. The device can be ~cce~e~l through a notch 334 formed in the second opposable component 304.
In use, a blood sample is applied to the porous pad 322 for separating the cellular components of blood. The liquid portion of the blood 10 sample then migrates through the chromatographic mP-lium 310; at that point, the first and second opposable components 302 and 304 are brought into opposition, and a reagent in the applicator 308 is applied to the chromatographic mto-linm 310 and migrates through the chromatographic m-o-linm 310 in the opposite direction from the flow of the liquid portion of the blood sample through the 15 chromatographic m(~ lm 310, thus reversing the flow. The reversal of flow is driven by the absorber 306.
This embodiment is particularly suitable for the pelrollllallce of serological assays to detect antibodies in blood samples. For example, if the analyte to be ~1etecte~1 is human antibody to the bacterium Helicobacter pylori,believed to be the cause of stomach ulcers, a blood sample suspected of cont~inin~ the antibody can be applied to the porous pad 322 to s~al~Lt: the cellular components of the blood sample from the liquid portion of the blood sample. The liquid portion of the blood sample then migrates from the porous pad 322 to the chromatographic mP-1inm 310. The detection zone 316 can contain immobilized H. pylori antigen, so that any antibody specific for H. pylori antigen binds at the ~l~t~ction zone. The applicator 308 then contains a labeledantibody that binds human immllnoglobulin G antibody, such as a gold-labeled goat anti-human immlmoglobulin G antibody, in resolubilizable form. The contents of the applicator 308 are resolubilized by the addition of an aqueous liquid to the applicator 308. When the first and second opposable components 302 and 304 are brought into opposition, the applicator 308 is brought into contact with the chromatographic mPrlillm 310 to apply the labeled anti-human CA 022l72l0 l997-l0-02 W 096/35952 PCTrUS96/06398 IgG antibody to the chromatographic medium 310. The absorber 306 then causes the labeled anti-human IgG antibody to migrate through the chromatographic medium 310 in a direction opposite to the flow of the liquid portion of the blood sample through the cl~olllatographic mtof~ m 310. Any anti-H. pylori antibody bound at the detection zone 316 then becomes labeled. If gold-labeled antibody is used, the presence of anti-H. pylori antibody can be ~let~cte~l visually. Thereverse flow, driven by the absorber 306, acts as a wash to remove other antibody present in the sample that is not specific for H. pylori antigen and is not bound at the detection zone 316 but would otherwise react with the labeled anti-human IgG antibody and give a background. The use of bidirectional flow therefore reduces the background and increases the sel~iLiviLy and reliability of the test.
These arr~ngçments are exemplary and are not exhaustive; other arrangements of both unidirectional and bidirectional assay devices according tothe present invention incorporating the porous pad for binding the cellular components of blood are also within the scope of the present invention. These arrangements can include a number of elements.
For example, in a number of devices according to the present invention, absorbers are in operable contact with one end of the chromatographicmedium, typically the end opposite from the end with which contact is made with the pad of porous m~teri~l. The absorbers can be made of any bibulous material that will hold a liquid sufficiently so liquid can be drawn through the chlolllat~graphic m~-lillm and accl-m~ tPcl in the absorber. Typical materials for the absorbers include, but are not limited to, filter paper.
Additionally, the devices can include one or more conductors.
Con~ ctors can serve as a bridge between the pad of porous m~teri~l and the chromatographic m.ot1illm wherefore CO.~ i..g the means for withdrawing the - liquid portion of blood from the porous material. These conductors are prepared of hydrophilic media that pass liquids without substantially absorbing them.
Such materials are well-known in the art. Cellulose and cellulose derivatives can be used.

W 096/35952 PCTrUS96/06398 In devices according to the present invention that employ opposable components, the bodies of the opposable components are preferably made of l~min~t-o~ cardboard that is sufficiently impervious to moisture to contain the liquids involved in the p~lr~ e of the assay carried out by the 5 device. Other cellulose-based materials, such as paperboard or solid bleached sulfite (SBS) can also be used. ~lt~rn~tively, the bodies of the opposable components can be made of plastic that is hll~el ~/ious to moisture. A suitable plastic is a polycarbonate plastic such as Lexan~-The opposable components are joined by a hinge, preferably made 10 of a material impermeable to liquids, such as a plastic that can be comp~tibly joined with or is the same as a material used for the first and second opposablecomponents.
A version particularly adapted for the performance of bidirectional assays can cc,lllplise:
(1) a first opposable component in~ ling:
(a) a first porous ~ ldLulg matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood, and (b) a second porous matrix in(~ ing a membrane for chromatographic separation in operable contact with the first porous separating 20 matrix that permits the liquid portion of the blood to flow in a first direction by capillary action or cln..lllalographic separation through the second porous matrix, and (2) a second opposable component ~tt~rh~hle to the first opposable component such that the first and second opposable components can be brought into opposition to L.al~irel a reagent from the second opposable component to the first opposable component by ~ s~ule so bringing the first and second opposable components into opposition causes the reagent LL~ul~rell~d from the second opposable component to the first opposable component to migrate through the second porous matrix in a second direction opposite to the first direction.
Typically, in this version, the membrane for chromatographic sepald~ion includes therein a capture zone for binding an analyte and the reagent -W 096/35952 PCTrUS96/06398 transferred from the second opposable component to the first opposable component is a labeled specific binding partner for the analyte.
A method for pclrOll~ g an assay for cletecting and/or ~leterminin~ at least one analyte in the liquid portion of a blood sample using this 5 version can comprise the steps of:
(1) applying a sample of blood to the first porous separating matrix on the first opposable component of the device;
(2) allowing the blood sample to flow through the first porous sel)dldLillg matrix to separate the liquid portion of the blood sample from the 10 cellular components of the blood sample;
(3) allowing the liquid portion of the blood sample to flow through the membrane for chromatographic separation in the first direction;
(4) bringing the first and second opposable components into opposition to Lldl~rcl the labeled specific binding partner for the analyte from the 15 second opposable colll~ollenl to the first opposable component by pressure; and (5) allowing the labeled specific binding partner for the analyte to flow through the membrane for chromatographic separation in the second direction so that an assay is performed in the second matrix, the assay being performed by binding the labeled specific binding partner to the capture zone of20 the second matrix to detect and/or dc~cllll~le the at least one analyte.
The description of the devices above is directed to assay devices that perform one assay at a time. However, assay devices according to the present invention can also be constructed that can perform multiple assays at the same time. The assays can be performed for the same analyte or dirrclclll 25 analytes. This allows the application of multiple blood samples to a single device with the performance of multiple assays.

II. DEVICES AND METHODS FOR OFF-BOARD PROCESSING
Another aspect of the present invention is a method for sc~aldlillg 30 the liquid portion of blood from the cellular components of blood by the pre-addition of a binder for the cellular components of blood to a sample of whole W 096/35952 PCTrUS96/06398 blood before the mi~Lule is applied to a device for sepalaLillg the liquid portion of blood from the cellular components of blood.
One such method comprises:
(1) adding a cross-linking substance for the cellular components of S blood to a sample of whole blood, the cross-linking substance being selected from the group consisting of a lectin, an anti-blood cell antibody, and a carbohydrate capable of aggregating blood cells;
(2) mixing the cross-linking substance and the blood sample to form a mixture of the cross-linking substance and the blood sample, or allowing 10 time for mixing to occur;
(3) applying the mixture of the cross-linking substance and the blood sample to a device for sepalaLillg the liquid portion of blood from the cellular components of blood, the device colllpli~ g:
(a) a pad of porous material permeable to the liquid portion 15 of blood but capable of trapping the cellular components of blood aggregated by the reaction between the cross-linking substance and the blood sample;
(b) a substrate supporting the pad; and (c) means, attached to the pad, for facilit~ting the flow of the liquid portion of the blood: (i) through interstices around the trapped cellular components of the blood and (ii) from the pad of porous material; and (4) allowing the liquid portion of the blood to flow through the pad to separate the liquid portion of the blood from the cellular collll~ont;llLs of the blood.
The separation of the liquid portion of blood from the cellular components of the blood bound to the binder occurs by flow through the pad without .signifir~nt hemolysis. This method differs from the methods described above in that the pad of porous material need not contain a cross-linking substance such as an antibody or a lectin; rather the pad acts as a filter to remove cellular components of blood aggregated by previous binding to the cross-linkingsubstance, with the binding occurring before the sample is applied to the pad.

W 096/35952 PCTrUS96/06398 Preferably, an anticoagulant is added with the cross-linking substance. A typical anticoagulant is EDTA or heparin, although other anticoagulants are known in the art.
Preferably, a concentration of cross-linking substance is used that r 5 is sufficient to cross-link subst~nti~lly all cellular element~ of blood.
The device for s~alaLil,g the liquid portion of blood from the cellular components of blood can be any of the allel,lalivt;s described above inSection I, with the difference that the pad of porous material acts as a filter for removing already aggl~ tr~l or aggregated cellular components of blood instead of providing means for aggll.~ n or aggregation of the cellular components.
The separated liquid portion of the blood can then be assayed for an analyte as described above, typically by an immlmochromatographic procedure. If the device used for s~al~ lg the liquid portions of blood from the cellular components of blood includes a chromatographic medium, as described above, the assay can be performed in the device; this is generally ~r~r~ d. Otherwise, the separated liquid portion of the blood can be drawn off for an assay on another device. These assays can be performed on assay devices such as those disclosed in copending U.S. Patent Application Serial No.
08/040,430 by Howard M. Chandler et al., entitled "Opposable-Flem~nt Chromatographic Assay Device," incorporated herein by l~,ferellce. These devices include both unidirectional and bidirectional assay devices.
AlLelllaliv~ly, instead of adding the cross-linking sllbst~nre to a sample of whole blood, a sample of blood can be added to a capillary tube coatedwith a cross-linking substance, with or without an anticoagulant. The cross-linking substance and anticoagulant, if present, are then allowed to dissolve inthe blood sample. The blood sample with the cross-linking sllbst~nre and anticoagulant dissolved therein is then applied to the device for separating the- liquid portion of blood from the cellular components of blood as described above. Again, the device acts as a filter for the aggl~ tr~l or aggregated bloodcells. An assay can be performed as described above.

This Z~ iv~ is shown generally in Figure 8. The blood sample 400 is added to the capillary tube 402 and, after mixing, the capillary tube is applied to the separation device 404.

ADVANTAGES OF THE INVENTION
The present invention provides a rapid, efficient, and simple means of sepald~ g blood cells from the liquid portion of blood for the performance of specific binding assay such as immlln~assays as well as other tests. In particular, the present invention provides an integrated device that incorporates both an assay element and means for s~al~Lh~g the liquid portions of blood from the cellular components of blood so that an analyte present in theliquid portions of blood can be assayed readily. This avoids the nPces~ity of a preliminary extraction of serum or plasma from blood with the intended n~ce~ity of safe disposal of the blood fractions. The use of an assay device according tothe present invention allows for convenient and safe disposal of used test devices.
Additionally, the improved device is capable of direct assay of a desired analyte when a whole blood sample is applied to the device.
Assay devices according to the present invention can perform a broad range of immlln~ says, in~ ing both sandwich and competitive immlln~assays. In particular, assay devices according to the present invention are suitable for the detection and/or .1~l~"..i~-~tion of both antigens and antibodies.
Although the present invention has been described with considerable detail, with reference to certain pl~rellcd variations thereof, other 25 versions and embo-limentc are possible. These versions include other arrangements of two-component devices that operate by the basic principles described herein. These versions include assay devices adapted for competitive immllnoassays as well as sandwich immllnoa~says in various arrangements. In particular, devices according to the present invention can be adapted to make use 30 of radial or cil~ f~l~--Lidl flow through a chromatographic m~-lillm rather than linear flow. Devices according to the present invention can also be adapted to ~elrollll multiple assays ~imlllt~nPously, with multiple second porous matrices, CA 022l72l0 l997-l0-02 W 096/35952 PCT~US96/06398 arranged circularly or like the spokes of a wheel, or in other arrangements.
Although devices according to the present invention are particularly adapted forthe ~e~ lion of the liquid portion of blood from the cellular components of blood and the pclrollllance of assays on the liquid portion of blood, devices according to the present invention can also be used for the removal of blood cells from other body fluids that may contain them, such as cerebrospinal fluid, and for assays on such fluids after the removal of blood cells from them. Devices according to the present invention can also be adapted to perform other assays, such as enzyme assays and colorimetric assays.
The present invention further encompasses variations in which the two components of the device are not held in a permanently fixed arrangement, but can be separated and brought together to perform the assay, such as by electrical or m~gnPtic forces or by using a separable fastener such as a hook-and-eye fabric, for example Velcro~. Additionally, the present invention can also encompass devices having three components in a folding arrangement.
Therefore, the scope of the invention is ~iett-.rmin~A by the following claims.

Claims (46)

1. A device for separating the liquid portion of blood from the cellular components of blood comprising:
(a) a pad of porous material permeable to the liquid portion of blood but capable of trapping the cellular components of blood;
(b) a substrate supporting the pad: and (c) means, attached to the pad, for facilitating the flow of the liquid portion of the blood; through interstices around the trapped cellular componentsof the blood and (ii) from the pad of porous material;
whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad to the means for facilitating the flow without significant hemolysis.
2. The device of claim 1 wherein the pad of porous material contains a binder for the cellular components of blood.
3. The device of claim 1 wherein the pad is impregnated with a carbohydrate capable of aggregating blood cells.
4. The device of claim 1 wherein the pad of porous material includes two sectors:
(i) a first sector permeable to both the liquid portion of blood and the cellular components of blood; and (ii) a second sector permeable to the liquid portion of blood but capable of binding the cellular components of blood.
5. The device of claim 1 wherein the pad of porous material permeable to the liquid portion of blood but capable of trapping the cellular components of blood includes therein an asymmetric membrane with a first surfaceand a second surface, the membrane having a gradient of pore sizes such that the pore size decreases from the first surface to the second surface, the asymmetricmembrane capable of trapping the cellular components of blood within it and allowing the liquid components of blood to pass through.
6. The device of claim 1 wherein the means, attached to the pad, for facilitating the flow of the liquid portion of the blood includes therein a membrane for chromatographic separation.
7. A device for separating the liquid portion of blood from the cellular components of blood consisting essentially of:
(a) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (b) a second porous matrix in operable contact with the first porous separating matrix that permits the liquid portion of the blood to flow by capillary action through the second porous matrix;
whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the first and second matrices from the firstmatrix to the second matrix without significant hemolysis.
8. A device for separating the liquid portion of blood from the cellular components of blood consisting essentially of:
(a) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood;
(b) a second porous separating matrix in operable contact with the first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (e) a third porous matrix in operable contact with the second porous separating matrix that permits the liquid portion of the blood to flow by capillary action or chromatographic separation through the second porous matrix;
whereby the separation of the liquid portion of the blood from the cellular components of the blood occurs by flow through the first and second porous separating matrices from the first matrix to the second matrix without significant hemolysis.
9. The device of claim 8 wherein either the first porous matrix, the second porous matrix, or both the first and second porous matrices contain a binder for the cellular components of blood.
10. The device of claim 8 wherein either the first porous matrix, the second porous matrix, or both the first and second porous matrices are impregnated with a carbohydrate capable of aggregating blood cells.
11. The device of claim 8 wherein either the first porous matrix, the second porous matrix, or both the first and second porous matrices include two sectors:
(i) a first sector permeable to both the liquid portion of blood and the cellular components of blood; and (ii) a second sector permeable to the liquid portion of blood but capable of binding the cellular components of blood.
12. The device of claim 8 wherein either the first porous matrix, the second porous matrix, or both the first and second porous matrices are asymmetric membranes with a first surface and a second surface, the membrane having a gradient of pore sizes such that the pore size decreases from the first surface to the second surface, the asymmetric membrane capable of trapping the cellular components of blood within it and allowing the liquid components of blood to pass through.
13. The device of claim 8 wherein the third matrix includes a membrane for chromatographic separation,
14. A device for separating the liquid portion of blood from the cellular components of blood comprising:
(a) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (b) at least two second porous matrices, each second porous matrix in operable contact with the first porous separating matrix that permits the liquid portion of the blood to flow by capillary action or chromatographic separation through the second porous matrix;
whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the first and second matrices without significant hemolysis.
15. The device of claim 14 wherein each second matrix includes a membrane for chromatographic separation.
16. A device for separating the liquid portion of blood from the cellular components of blood comprising:
(a) a first opposable component including:
(i) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (ii) a second porous matrix in operable contact with the first porous separating matrix that permits the liquid portion of the blood to flow bycapillary action or chromatographic separation through the second porous matrix;and (b) a second opposable component attachable to the first opposable component such that the first and second opposable components can be brought into opposition to transfer fluid from one of the opposable components to the other by pressure;
whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the first and second matrices of the first opposable component without significant hemolysis.
17. The device of claim 16 wherein the second opposable component includes a sample application zone.
18. The device of claim 17 wherein the sample application zone includes a specific binding partner labeled with a detectable label, the specific binding partner having specific binding affinity for at least one component selected from the analyte and a specific binding partner for the analyte in a form that can be resolubilized by the addition of an aqueous sample to the sample preparation zone.
19. The device of claim 16 wherein at least one of the first and second opposable components further includes a sample application zone including a specific binding partner labeled with a detectable label, the specific binding partner having specific binding affinity for at least one component selected from the analyte and a specific binding partner for the analyte in a form that can be resolubilized by the addition of an aqueous sample to the sample application zone.
20. A device for separating the liquid portion of blood from the cellular components of blood comprising:
(a) a first opposable component including:
(i) a first porous separating matrix permeable to the liquid portion of blood but capable of trapping the cellular components of blood; and (ii) a second porous matrix including a membrane for chromatographic separation in operable contact with the first porous separating matrix that permits the liquid portion of the blood to flow in a first direction by capillary action or chromatographic separation through the second porous matrix;and (b) a second opposable component attachable to the first opposable component such that the first and second opposable components can be brought into opposition to transfer a reagent from the second opposable component to the first opposable component by pressure so bringing the first and second opposable components into opposition causes the reagent transferred from the second opposable component to the first opposable component to migrate through the second porous matrix in a second direction opposite to the first direction;
whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the first and second matrices of the first opposable component without significant hemolysis.
21. The device of claim 20 wherein the membrane for chromatographic separation includes therein a capture zone for binding an analyte and the reagent transferred from the second opposable component to the first opposable component is a labeled specific binding partner for the analyte.
22. The device of any of claims 7, 14, or 16 wherein the first porous separating matrix contains a binder for the cellular components of blood.
23. The device of any of claims 2 or 9 wherein the binder is an anti-blood cell antibody.
24. The device of claim 22 wherein the binder is an anti-blood cell antibody.
25. The device of claim 23 wherein the anti-blood cell antibody is an anti-erythrocyte antibody.
26. The device of claim 24 wherein the anti-blood cell antibody is an anti-erythrocyte antibody.
27. The device of any of claims 2 or 9 wherein the binder is a lectin.
28. The device of claim 27 wherein the lectin is selected from the group consisting of concanavalin A, abrin, phytohaemagglutinin, limulin, and the lectins produced by the following species: Agaricus bisporus, Anguilla anguilla,Arachis hypogaea, Bandeiraea simplicifolia, Bauhinia purpurea, Caragana arborescens, Cicer arietinum, Codium fragile, Datura stramonium, Dolichos biflorus, Erythrina corallodendron, Erythrina cristagalli, Euonymus europaeus, Glycine max, Helix aspersa, Helix pomatia, Lathyrus odoratus, Lens culinaris, Lycopersicon esculentum, Maclura pomifera, Momordica charantia, Mycoplasma gallisepticum, Naja mocambique, Naja kaouthia, Perseau americana, Phaseolus coccineus, Phaseolus limensis, Phaseolus vulgaris. Phytolacca americana, Pisum sativum, Pseudomonas aeruginosa, Psophocarpus tetragonolobus, Ptilota plumosa, Ricinus communis, Robinia pseudoacacia, Sambucus nigra, Solanum tuberosum, Sophora japonica, Tetragonolobus purpureas, Triticum vulgaris, Ulex europaeus, Vicia faba, Vicia sativa, Vicia villosa, Vigna radiata, Viscum album, and Wisteria floribunda.
29. The device of claim 22 wherein the binder is a lectin.
30. The device of claim 29 wherein the lectin is selected from the group consisting of concanavalin A, abrin, phytohaemagglutinin, limulin, and thelectins produced by the following species: Agaricus bisporus, Anguilla anguilla,Arachis hypogaea, Bandeiraea simplicifolia, Bauhinia purpurea, Caragana arborescens, Cicer arietinum, Codium fragile, Datura stramonium, Dolichos biflorus, Erythrina corallodendron, Erythrina cristagalli, Euonymus europaeus, Glycine max, Helix aspersa, Helix pomatia, Lathyrus odoratus, Lens culinaris.
Lycopersicon esculentum. Maclura pomifera, Momordica charanria, Mycoplasma gallisepticum, Naja mocambique, Naja kaouthia, Perseau americana, Phasealus coccineus, Phaseolus limensis, Phaseolus vulgaris, Phytolacca americana, Pisum sativum, Pseudomonas aeruginosa, Psophocarpus tetragonolobus, Ptilota plumosa, Ricinus communis, Robinia pseudoacacia, Sambucus nigra, Solonum tuberosum, Sophora japonica, Tetragonolobus purpureas, Triticum vulgaris, Ulex europaeus, Vicia faba, Vicia sativa, Vicia villosa, Vigna radiata, Viscum album, and Wisteria floribunda.
31. The device of any of claims 7, 14, or 16 wherein the first porous separating matrix is impregnated with a carbohydrate capable of aggregating blood cells.
32. The device of claim 31 wherein the carbohydrate is selected from the group consisting of mannitol, sorbitol, inositol, b-D-glucose, a-D-glucose, D(+)xylose, D(+)mannose D(-)arabinose, L(+)arabinose, D(+)galactose, L(-)xylose,D-glucoheptose, L-lyxose, lactose, maltose, and sucrose.
33. The device of claim 32 wherein the carbohydrate is mannitol.
34. The device of any of claims 3 or 10 wherein the carbohydrate is selected from the group consisting of mannitol, sorbitol, inositol, b-D-glucose,a-D-glucose, D(+)xylose, D(+)mannose, D(-)arabinose, L(+)arabinose, D(+)galactose, L(-)xylose, D-glucoheptose, L-lyxose, lactose, maltose, and sucrose.
35. The device of claim 34 wherein the carbohydrate is mannitol.
36. The device of any of claims 7 or 16 wherein the second matrix includes a membrane for chromatographic separation.
37. The device of claim 36 wherein the membrane for chromatographic separation has a capture zone thereon for binding a member of a specific binding pair.
38, The device of any of claims 6, 13, or 15 wherein the membrane for chromatographic separation has a capture zone thereon for binding a member of a specific binding pair.
39. The device of any of claims 7 or 16 wherein the first separating matrix is an asymmetric membrane with a first surface and a second surface, the membrane having a gradient of pore sizes such that the port size decreases from the first surface to the second surface, the asymmetric membrane capable of trapping the cellular components of blood within it and allowing the liquid components of blood to pass through,
40. the device of any of claims 7 or 16 wherein the first separating matrix includes two sectors:
(i) a first sector permeable to both the liquid portion of blood and the cellular components of blood; and (ii) a second sector permeable to the liquid portion of blood but capable of binding the cellular components of blood.
41. A method for separating the liquid portion of blood from the cellular components of blood comprising the steps of:
(a) adding a cross-linking substance for the cellular components of blood to a sample of whole blood, the cross linking substance being selected from the group consisting of a lectin, an anti-blood cell antibody, and a carbohydrate capable of agglutinating blood cells;
(b) mixing the cross-linking substance and the blood sample to form a mixture of the cross-linking substance and the blood sample;
(c) applying the mixture of the cross-linking substance and the blood sample to a device for separating the liquid portion of blood from the cellular components of blood, the device comprising:
(i) a pad of porous material permeable to the liquid portion of blood but capable of trapping the cellular components of blood aggregated by thereaction between the cross-linking substance and the blood sample;
(ii) a substrate supporting the pad; and (iii) means, attached to the pad, for facilitating the flow of the liquid portion of the blood: through interstices around the trapped cellular components of the blood and (2) from the pad of porous material, whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad without significant hemolysis; and (d) allowing the liquid portion of the blood to flow through the pad to separate the liquid portion of the blood from the cellular components of the blood.
42. The method of claim 41 further comprising adding an anticoagulant together with the cross-linking substance.
43. A method for separating the liquid portion of blood from the cellular components of blood comprising the steps of:
(a) adding a sample of blood to a capillary tube coated with a cross-linking substance, the cross-linking substance being selected from the group consisting of a lectin, an anti-blood cell antibody, and a carbohydrate capable of aggregating blood cells;
(b) allowing the cross-linking substance to dissolve in the blood sample to form a mixture of the cross-linking substance and the blood sample;
(c) applying the mixture of the cross-linking substance and the blood sample to a device for separating the liquid portion of blood from the cellular components of blood, the device comprising:
(i) a pad of porous material permeable to the liquid portion of blood but capable of trapping the cellular components of blood aggregated by thereaction between the cross-linking substance and the blood sample;
(ii) a substrate supporting the pad; and (iii) means, attached to the pad, for facilitating the flow of the liquid portion of the blood; (1) through interstices around the trapped cellularcomponents of the blood and (2) from the pad of porous material, whereby the separation of the liquid portion of blood from the cellular components of the blood occurs by flow through the pad without significant hemolysis; and (d) allowing the liquid portion of the blood to flow through the pad to separate the liquid portion of the blood from the cellular components of the blood
44. The method of claim 43 wherein the capillary tube is also coated with an anticoagulant together with the cross-linking substance and the anticoagulant is allowed to dissolve in the blood sample.
45. The method of any of claims 42 or 44 wherein the anticoagulant is selected from the group consisting of EDTA and heparin.
46. The method of any of claims 41 or 43 wherein a concentration of cross-linking substance is used that is sufficient to cross-link substantially all cellular elements of blood.
CA002217210A 1995-05-09 1996-05-07 Devices and methods for separating cellular components of blood from liquid portion of blood Abandoned CA2217210A1 (en)

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Families Citing this family (99)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR100241928B1 (en) * 1997-03-31 2000-03-02 박종근 Determination device in which the electrode is integrally formed on the porous thin film and the quantification method using the same
AU738143B2 (en) * 1997-10-07 2001-09-13 Neogen Corporation Assay device for determining analytes in a liquid dairy product
GB9821526D0 (en) * 1998-10-02 1998-11-25 Genosis Inc Capture assay
US6036659A (en) 1998-10-09 2000-03-14 Flexsite Diagnostics, Inc. Collection device for biological samples and methods of use
US6511814B1 (en) * 1999-03-26 2003-01-28 Idexx Laboratories, Inc. Method and device for detecting analytes in fluids
US6551842B1 (en) 1999-03-26 2003-04-22 Idexx Laboratories, Inc. Method and device for detecting analytes in fluids
GB9929272D0 (en) * 1999-12-10 2000-02-02 Diagnology Limited Assay
US6316205B1 (en) * 2000-01-28 2001-11-13 Genelabs Diagnostics Pte Ltd. Assay devices and methods of analyte detection
JP4681198B2 (en) * 2000-03-17 2011-05-11 株式会社三和化学研究所 Test paper
KR100513213B1 (en) * 2000-06-01 2005-09-08 마츠시타 덴끼 산교 가부시키가이샤 Biosensor and blood component analyzing method
DE60142394D1 (en) * 2000-06-28 2010-07-29 Panasonic Corp BIOZENSOR
US6612447B1 (en) * 2000-07-24 2003-09-02 Baxter International Inc. Blood collection systems and filters using a porous membrane element
US20020036170A1 (en) * 2000-08-11 2002-03-28 Harvey Michael A. Lateral flower plasma separation device
CN1302285C (en) * 2001-01-12 2007-02-28 东丽株式会社 Selectively hybridizable substance immobilization fiber, fiber array comprising bundle of such fibers, selective hybridizing method, device therefor and base
CN100437114C (en) 2001-04-12 2008-11-26 爱科来株式会社 Specimen analyzing implement
US8101077B2 (en) * 2001-05-07 2012-01-24 Sivaprasad Sukavaneshvar Device for separating platelets from fluid suspensions
US7011852B2 (en) * 2001-05-07 2006-03-14 Hemogenesis, Llc Separation of platelets from whole blood for use as a healant
KR20010085007A (en) * 2001-07-12 2001-09-07 김희태 A semi-quantitative membrane immunochromatographe and a method for the preparation thereof
CA2488933C (en) * 2002-04-23 2016-11-01 Home Access Health Corporation Quantitative analysis of a biological sample of unknown quantity
US7569184B2 (en) * 2002-04-23 2009-08-04 Home Access Health Corporation Quantitative analysis of a biological sample of unknown quantity
US7611670B2 (en) * 2002-04-23 2009-11-03 Home Access Health Corporation Quantitative analysis of a biological sample of unknown quantity
US20040048359A1 (en) * 2002-07-12 2004-03-11 Schmeling William R. Test strips moveable by magnetic fields
JP4339570B2 (en) * 2002-10-25 2009-10-07 株式会社エスアールエル Blood sample carrier component
CN101819198B (en) * 2003-03-10 2012-07-18 爱科来株式会社 Blood cell separation membrane
US8454566B2 (en) * 2003-07-10 2013-06-04 Medtronic Minimed, Inc. Methods and compositions for the inhibition of biofilms on medical devices
US20050025823A1 (en) * 2003-07-29 2005-02-03 Fong Andy A.T. Methods of use of herbal compositions
US7625721B2 (en) 2004-02-03 2009-12-01 Polymer Technology Systems, Inc. Non-precipitating bodily fluid analysis system
US8465696B2 (en) * 2004-02-03 2013-06-18 Polymer Technology Systems, Inc. Dry test strip with controlled flow and method of manufacturing same
US20060062688A1 (en) * 2004-02-03 2006-03-23 Polymer Technology Systems, Inc. Bodily fluid analysis system
CN100359314C (en) * 2004-02-17 2008-01-02 陈格 Method and device for drying and preserving liquid sample
ATE489625T1 (en) * 2004-06-07 2010-12-15 Denka Seiken Kk CHROMATOGRAPHIC DETECTION DEVICE
WO2006004039A1 (en) * 2004-07-02 2006-01-12 Sanwa Kagaku Kenkyusho Co., Ltd. Immunoassay apparatus and method
US20060019406A1 (en) * 2004-07-23 2006-01-26 Ning Wei Lateral flow device for the detection of large pathogens
WO2006017703A1 (en) * 2004-08-05 2006-02-16 Akers Biosciences, Inc. Blood separator and method of separating a fluid fraction from whole blood
WO2006023679A1 (en) * 2004-08-17 2006-03-02 Polymer Technology Systems, Inc. Apparatus and method for manufacturing bodily fluid test strip
US20060068500A1 (en) * 2004-09-28 2006-03-30 Kimberly-Clark Worldwide, Inc. Detecting yeast infections using a lateral flow assay
US7442557B1 (en) 2004-10-22 2008-10-28 Idexx Laboratories, Inc. Bi-directional flow assay device with reagent bridge
JP2006133143A (en) * 2004-11-08 2006-05-25 Japan Vilene Co Ltd Blood plasma or blood serum holding material and blood separation tool
NL1027737C2 (en) * 2004-12-14 2006-06-16 Teststrip B V Chromatographic analysis device and test kit.
ATE490462T1 (en) * 2005-04-08 2010-12-15 Medical Discovery Partners Llc METHOD FOR ENRICHING SUBPOPULATIONS OF RARE CELLS FROM THE BLOOD
AU2006321289B2 (en) * 2005-05-23 2011-12-08 Phadia Ab Two step lateral flow assay methods and devices
WO2007000048A1 (en) * 2005-06-28 2007-01-04 Zbx Corporation Membrane array and analytical device
US7618810B2 (en) * 2005-12-14 2009-11-17 Kimberly-Clark Worldwide, Inc. Metering strip and method for lateral flow assay devices
US8377379B2 (en) * 2006-12-15 2013-02-19 Kimberly-Clark Worldwide, Inc. Lateral flow assay device
JP2008249606A (en) * 2007-03-30 2008-10-16 Sysmex Corp Testing implement for chromatography
JP4990692B2 (en) * 2007-06-22 2012-08-01 株式会社ビーエル Immunochromatographic assay and kit
US8535617B2 (en) * 2007-11-30 2013-09-17 Kimberly-Clark Worldwide, Inc. Blood cell barrier for a lateral flow device
WO2009075709A1 (en) * 2007-12-12 2009-06-18 Micropoint Bioscience Inc Rapid and efficient filtering whole blood in a capillary flow device
US9968931B2 (en) * 2007-12-12 2018-05-15 Nan Zhang Rapid and efficient filtering whole blood in capillary flow device
US8613214B2 (en) * 2008-01-09 2013-12-24 Orono Spectral Solutions, Inc. Apparatus and method for determining analyte content in a fluid
US8393198B2 (en) * 2008-01-09 2013-03-12 OronoSpectral Solutions, Inc. Apparatus and method for determining analyte content in a fluid
JP4428670B2 (en) 2008-03-06 2010-03-10 Ｔａｎａｋａホールディングス株式会社 Immunological assay, kit and developing solvent
DK2260300T3 (en) 2008-03-07 2014-01-13 Advanced Microdevices Pvt Ltd Method and Device for Particle Removal and Preparation of Drops for Qualitative and Quantitative Bioassay
EP2257818B1 (en) * 2008-03-27 2017-05-10 President and Fellows of Harvard College Cotton thread as a low-cost multi-assay diagnostic platform
JP2009236685A (en) * 2008-03-27 2009-10-15 Sysmex Corp Chromatographic test device
JP5117928B2 (en) 2008-05-27 2013-01-16 富士フイルム株式会社 Immunochromatographic device
EP2326419B1 (en) 2008-06-29 2021-04-07 Realbio Technologies Ltd. Liquid transfer device particularly useful as a capturing device in a biological assay process
KR101032691B1 (en) * 2009-04-17 2011-05-06 (주)디지탈옵틱 Biosensor for the use of diagnosis that prompt blood separation is possible
CN102414561A (en) * 2009-04-28 2012-04-11 创新实验室技术公司 Lateral-flow immuno-chromatographic assay devices
US9475914B2 (en) * 2010-01-08 2016-10-25 University Of Tasmania Porous polymer monoliths, processes for preparation and use thereof
US9816979B2 (en) * 2010-07-27 2017-11-14 Northwestern University Devices and methods for filtering blood plasma
US8956859B1 (en) 2010-08-13 2015-02-17 Aviex Technologies Llc Compositions and methods for determining successful immunization by one or more vaccines
CN102375055A (en) * 2010-08-19 2012-03-14 中国人民解放军军事医学科学院微生物流行病研究所 Multiplex detection immune chromatography chip
EP2785429A4 (en) 2011-11-30 2015-10-28 Wellstat Diagnostics Llc Filtration module
CN105833925B (en) * 2011-12-22 2018-11-13 瑞尔比奥技术有限公司 sequential lateral flow capillary device for analyte determination
AU2013203281A1 (en) 2012-02-03 2013-08-22 Axxin Pty Ltd Nucleic acid amplification and detection apparatus and method
US9880164B2 (en) 2012-03-30 2018-01-30 Sekisui Medical Co., Ltd. Immunochromatographic test strip and detection method using immunochromatography for detecting object in red blood cell-containing sample
US9081001B2 (en) 2012-05-15 2015-07-14 Wellstat Diagnostics, Llc Diagnostic systems and instruments
US9213043B2 (en) 2012-05-15 2015-12-15 Wellstat Diagnostics, Llc Clinical diagnostic system including instrument and cartridge
US9625465B2 (en) 2012-05-15 2017-04-18 Defined Diagnostics, Llc Clinical diagnostic systems
US10048262B2 (en) 2012-06-13 2018-08-14 Asahi Kasei Kabushiki Kaisha Method for detecting specific substance in milk
EP2883048A1 (en) * 2012-08-08 2015-06-17 Koninklijke Philips N.V. Plasma separation using a drop of blood
US9285336B2 (en) * 2012-08-09 2016-03-15 The Board Of Trustees Of The Leland Stanford Junior University Sensing platform for quantum transduction of chemical information
US10248765B1 (en) 2012-12-05 2019-04-02 Theranos Ip Company, Llc Systems, devices, and methods for bodily fluid sample collection, transport, and handling
CN111366442A (en) * 2013-03-15 2020-07-03 赛拉诺斯知识产权有限责任公司 Method and apparatus for sample collection and sample separation
US10101293B2 (en) 2013-08-09 2018-10-16 The Board Of Trustees Of The Leland Stanford Junio Sensing platform for transduction of information
CA2953755C (en) 2014-07-03 2022-05-03 Abionic Sa Capsule for rapid molecular quantification of a fluid sample such as whole blood
WO2016073415A2 (en) 2014-11-04 2016-05-12 Wainamics, Inc. Microscale plasma separator
ES2809600T3 (en) 2014-11-14 2021-03-04 Axxin Pty Ltd Biological sample collection and storage set
EP3294133B1 (en) * 2015-05-14 2019-09-04 General Electric Company Method and kit for separation and collection of plasma
US11198122B2 (en) 2015-07-17 2021-12-14 Axxin Pty Ltd Diagnostic test assembly, apparatus, method
US10371606B2 (en) 2015-07-21 2019-08-06 Theraos IP Company, LLC Bodily fluid sample collection and transport
WO2017044888A1 (en) 2015-09-09 2017-03-16 Theranos, Inc. Methods and devices for sample collection and sample separation
CN105445454B (en) * 2015-11-20 2016-11-30 广州瑞博奥生物科技有限公司 A kind of can be quantitative device for immunochromatography
US10983119B2 (en) * 2016-01-25 2021-04-20 General Electric Company Device for rapid diagnostic tests to detect antigens with improved sensitivity
WO2017189854A1 (en) 2016-04-28 2017-11-02 The Board Of Trustees Of The Leland Stanford Junior University Mesoscale system feedback-induced dissipation and noise suppression
JP6366025B2 (en) * 2016-10-03 2018-08-01 セルスペクト株式会社 Plasma separator and plasma separation method
GB201703383D0 (en) 2017-03-02 2017-04-19 Gargle Tech Ltd Testing for particulates
US11857966B1 (en) 2017-03-15 2024-01-02 Labrador Diagnostics Llc Methods and devices for sample collection and sample separation
WO2018175169A1 (en) 2017-03-20 2018-09-27 Wainamics, Inc. Small volume self-metered blood separation device
BR112020006004A2 (en) 2017-09-27 2020-10-06 Axxin Pty Ltd diagnostic test system, assembly and method
EP4205851A3 (en) 2018-09-05 2023-10-04 Hero Scientific Ltd Testing for particulates
CN109541215A (en) * 2018-12-20 2019-03-29 中国人民解放军陆军军医大学 A kind of lesion detection object and the preparation method and application thereof
US11389765B2 (en) * 2019-01-09 2022-07-19 Lawrence Livermore National Security, Llc Hierarchical triply periodic minimal surface structures as heat exchangers and reactors
US11885568B2 (en) 2019-01-09 2024-01-30 Lawrence Livermore National Security, Llc Systems and methods for periodic nodal surface based reactors, distributors, contactors and heat exchangers
EP3983127A4 (en) * 2019-06-12 2022-07-20 Siemens Healthcare Diagnostics, Inc. Plasma separation and sample metering device and kits and methods of use related thereto
CN111721880B (en) * 2020-07-02 2023-01-10 江中药业股份有限公司 Method for establishing fingerprint of sarcandra glabra by using double-column tandem HPLC-MS
WO2022149135A2 (en) 2021-01-06 2022-07-14 Hero Scientific Ltd. Filtration sampling devices
WO2023039536A1 (en) * 2021-09-10 2023-03-16 Quidel Corporation Agglutinating agents, devices and methods for whole blood assays

Family Cites Families (62)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3768978A (en) * 1971-03-01 1973-10-30 Hamilton Co Disposable pipette
US3723064A (en) * 1971-07-26 1973-03-27 L Liotta Method and device for determining the concentration of a material in a liquid
BE792129A (en) 1971-12-06 1973-05-30 Takeda Chemical Industries Ltd PYRIMIDO (4,5-D) DERIVATIVES PYRIDAZINE AND THEIR PRODUCTION PROCESS
US3902964A (en) * 1971-12-13 1975-09-02 U S Medical Research And Dev I Method of and apparatus for chemically separating plasma or serum from formed elements of blood
CA1054034A (en) 1975-06-20 1979-05-08 Barbara J. Bruschi Multilayer analytical element
US3990850A (en) * 1976-01-06 1976-11-09 Akzona Incorporated Diagnostic test card
US4110079A (en) 1977-06-09 1978-08-29 Eastman Kodak Company Analytical element for clinical analysis
US4160008A (en) * 1978-01-26 1979-07-03 Miles Laboratories, Inc. Multilayered test device for determining the presence of a liquid sample component, and method of use
JPS587332Y2 (en) 1978-06-06 1983-02-08 富士写真フイルム株式会社 Multilayer blood chemistry analysis material
DE2932973A1 (en) 1978-08-14 1980-02-28 Fuji Photo Film Co Ltd INTEGRAL MULTILAYERED MATERIAL FOR CHEMICAL ANALYSIS OF BLOOD
JPH0142041Y2 (en) * 1978-12-11 1989-12-11
US4288228A (en) 1979-01-31 1981-09-08 Technicon Instruments Corporation Whole blood analyses and diffusion apparatus therefor
US4271119A (en) * 1979-07-23 1981-06-02 Eastman Kodak Company Capillary transport device having connected transport zones
US4816224A (en) * 1980-08-05 1989-03-28 Boehringer Mannheim Gmbh Device for separating plasma or serum from whole blood and analyzing the same
DE3029579C2 (en) * 1980-08-05 1985-12-12 Boehringer Mannheim Gmbh, 6800 Mannheim Method and means for separating plasma or serum from whole blood
US4426451A (en) * 1981-01-28 1984-01-17 Eastman Kodak Company Multi-zoned reaction vessel having pressure-actuatable control means between zones
DE3323973A1 (en) * 1983-07-02 1985-01-03 Boehringer Mannheim Gmbh, 6800 Mannheim ERYTHROCYTE RETENTION SUBSTRATES
US4594327A (en) * 1983-11-02 1986-06-10 Syntex (U.S.A.) Inc. Assay method for whole blood samples
US4678757A (en) * 1985-04-11 1987-07-07 Smithkline Diagnostics, Inc. Device and method for whole blood separation and analysis
DE3523439A1 (en) 1985-06-29 1987-01-08 Boehringer Mannheim Gmbh TEST CARRIER
US4790979A (en) * 1986-08-29 1988-12-13 Technimed Corporation Test strip and fixture
DE3630999A1 (en) * 1986-09-12 1988-03-17 Boehringer Mannheim Gmbh MULTI-LAYER TEST CARRIER
US5135719A (en) * 1986-10-29 1992-08-04 Biotrack, Inc. Blood separation device comprising a filter and a capillary flow pathway exiting the filter
US4753776A (en) * 1986-10-29 1988-06-28 Biotrack, Inc. Blood separation device comprising a filter and a capillary flow pathway exiting the filter
DE3643516A1 (en) 1986-12-19 1988-06-30 Boehringer Mannheim Gmbh TEST CARRIER FOR THE ANALYTICAL DETERMINATION OF INGREDIENTS OF BODY LIQUIDS
DE3715485A1 (en) 1987-05-09 1988-11-17 Boehringer Mannheim Gmbh STABILIZATION OF SS GALACTOSIDASE
US4943522A (en) * 1987-06-01 1990-07-24 Quidel Lateral flow, non-bibulous membrane assay protocols
US4987085A (en) * 1987-06-22 1991-01-22 Chemtrak Inc. Blood filtering metering device
US5215886A (en) 1987-06-22 1993-06-01 Patel P Jivan HDL determination in whole blood
JPH0623759B2 (en) * 1987-07-08 1994-03-30 富士写真フイルム株式会社 Whole blood analysis element
US4883764A (en) 1987-07-20 1989-11-28 Kloepfer Mary A Blood test strip
US5209904A (en) 1987-12-23 1993-05-11 Abbott Laboratories Agglutination reaction device utilizing selectively impregnated porous material
NL8800796A (en) * 1988-03-29 1989-10-16 X Flow Bv METHOD FOR THE CHEMICAL ANALYSIS OF BODY FLUID COMPONENTS, AND A TESTING DEVICE AND TEST PACKAGE FOR SUCH ANALYSIS.
DE3814370A1 (en) 1988-04-28 1989-11-09 Boehringer Mannheim Gmbh TEST PROVIDER FOR THE ANALYSIS OF A SAMPLING FLUID, METHOD FOR CARRYING OUT SUCH ANALYSIS AND METHOD OF PRODUCTION
US4963325A (en) 1988-05-06 1990-10-16 Hygeia Sciences, Inc. Swab expressor immunoassay device
US5096809A (en) * 1988-07-25 1992-03-17 Pacific Biotech, Inc. Whole blood assays using porous membrane support devices
US5418129A (en) * 1989-01-30 1995-05-23 The Biomembrane Institute Blood treatment method
US5064541A (en) * 1989-04-07 1991-11-12 Abbott Laboratories Devices and methods for the collection of a predetermined volume of plasma or serum
US4933092A (en) * 1989-04-07 1990-06-12 Abbott Laboratories Methods and devices for the separation of plasma or serum from whole blood
NL8901090A (en) * 1989-04-28 1990-11-16 X Flow Bv METHOD FOR MANUFACTURING A MICROPOROUS MEMBRANE AND SUCH MEMBRANE
DE3922495A1 (en) * 1989-07-08 1991-01-17 Miles Inc ANALYSIS METHOD FOR SUBSTANCES FROM BIOLOGICAL FLUIDS, IN PARTICULAR FULLBLOOD
US5306623A (en) * 1989-08-28 1994-04-26 Lifescan, Inc. Visual blood glucose concentration test strip
EP0427534B1 (en) * 1989-11-08 1995-08-02 Chemtrak, Inc. Improvement in non-instrumental diagnostic assay distance determination
US5435970A (en) * 1989-12-18 1995-07-25 Environmental Diagnostics, Inc. Device for analysis for constituents in biological fluids
JP2618727B2 (en) 1990-01-19 1997-06-11 富士写真フイルム株式会社 Method for quantifying test components in whole blood
US5212060A (en) * 1990-04-27 1993-05-18 Genesis Labs, Inc. Dry test strip comprising a dextran barrier for excluding erythrocytes
DE4015589A1 (en) * 1990-05-15 1991-11-21 Boehringer Mannheim Gmbh DEVICE AND THE USE THEREOF FOR SEPARATING PLASMA FROM WHOLE BLOOD
US5118428A (en) * 1990-11-13 1992-06-02 Quidel Method to remove red blood cells from whole blood samples
US5468648A (en) * 1991-05-29 1995-11-21 Smithkline Diagnostics, Inc. Interrupted-flow assay device
US5877028A (en) * 1991-05-29 1999-03-02 Smithkline Diagnostics, Inc. Immunochromatographic assay device
EP0535485B1 (en) * 1991-10-03 1997-07-16 Bayer Corporation Device and method of separating and assaying whole blood
US5215102A (en) * 1991-10-25 1993-06-01 La Mina Ltd. Capillary blood antigen testing apparatus
JPH05188053A (en) * 1992-01-10 1993-07-27 Sanwa Kagaku Kenkyusho Co Ltd Instrument for separating serum or plasma component from blood
ES2151902T3 (en) * 1992-03-10 2001-01-16 Quidel Corp MEANS OF SEPARATION OF RED BALLOONS TO DOSAGE THEM BY SPECIFIC UNION.
GR1002549B (en) * 1992-05-12 1997-01-28 Lifescan Inc. Fluid conducting test strip with Transport Medium
CA2104976A1 (en) * 1992-09-02 1994-03-03 Stephan D. Daubney Separation of plasma or serum from whole blood using a red blood cell binding component and a polymer containing multiple cationic sites
US5354692A (en) * 1992-09-08 1994-10-11 Pacific Biotech, Inc. Analyte detection device including a hydrophobic barrier for improved fluid flow
US5460974A (en) * 1992-10-13 1995-10-24 Miles Inc. Method of assaying whole blood for HDL cholesterol
WO1994018559A1 (en) * 1993-02-11 1994-08-18 Radiometer Medical A/S Asymmetric membrane sensor
US5652148A (en) * 1993-04-20 1997-07-29 Actimed Laboratories, Inc. Method and apparatus for red blood cell separation
US5397479A (en) * 1993-04-26 1995-03-14 International Remote Imaging Systems, Inc. Composition and method for enrichment of white blood cells from whole human blood
US5441698A (en) * 1993-09-10 1995-08-15 Beckman Instruments, Inc. Bevel closure and device

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EP0832430B1 (en) 2006-07-26
JPH11505327A (en) 1999-05-18
US6069014A (en) 2000-05-30
US6197598B1 (en) 2001-03-06
US6008059A (en) 1999-12-28
CN1190464A (en) 1998-08-12
JP2005055451A (en) 2005-03-03
DE69636389D1 (en) 2006-09-07
DE69636389T2 (en) 2006-11-23
ATE334392T1 (en) 2006-08-15
JP3831403B2 (en) 2006-10-11
WO1996035952A1 (en) 1996-11-14
JP2006177970A (en) 2006-07-06
EP0832430A4 (en) 2000-12-06
ES2268706T3 (en) 2007-03-16
CN1155827C (en) 2004-06-30
EP0832430A1 (en) 1998-04-01
DK0832430T3 (en) 2006-11-20

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