CN105092543A - Portable fluorescence quantitative PCR detector - Google Patents
Portable fluorescence quantitative PCR detector Download PDFInfo
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- CN105092543A CN105092543A CN201410198605.6A CN201410198605A CN105092543A CN 105092543 A CN105092543 A CN 105092543A CN 201410198605 A CN201410198605 A CN 201410198605A CN 105092543 A CN105092543 A CN 105092543A
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Abstract
The invention discloses a portable fluorescence quantitative PCR detector, and belongs to the fields of biology and medical detection. The detector comprises an optical excitation unit, an optical detection unit, and a temperature control unit. The optical excitation unit comprises a plurality of light sources, an exciting light optical filter, and a holophote. The temperature control unit comprises a reaction tank, a heat cover, a heating/refrigerating sheet, a heat dissipating part, and a temperature control circuit. The optical detection unit comprises a plurality of light splitting sheets, an optical filter, an imaging sensor, and a detection circuit. The light given off by the light source goes through the exciting light optical filter and then is converted into exciting light with a specific wavelength; then the exciting light is reflected by the holophote and is projected into a PCR tube to excite the fluorescence substances in the reagent, thus the fluorescence substances will give off light, part of the light goes though the hole in the side surface of the reaction tank and enters the optical detection unit, the light is split and filtered in the optical detection unit, and finally the light form an image on the corresponding image sensor.
Description
Technical field
The present invention relates to a kind of Portable fluorescence quantitative PCR detector, belong to biology and medical science.
Background technology
PCR (PCR) is a kind of Protocols in Molecular Biology, for amplifying the specific DNA fragmentation of amplification.Real-Time Fluorescent Quantitative PCR Technique, refers to and add fluorophor in PCR reaction system, utilizes fluorescence signal to accumulate the whole PCR process of Real-Time Monitoring, finally by typical curve, unknown template is carried out to the method for quantitative test.
At present, fluorescent quantitative PCR detector on the market, difference maximum between each product is optical excitation and detection module, and these products mainly have the following disadvantages:
1. some fluorescent quantitative PCR detector uses photodiode as detector, each photodiode once can only detect a sample, so need to scan all samples by the mode of mechanical scanning, the existence of these mechanical hook-ups adds volume and lower power consumption reliability, the more important thing is that different sample has detection time poor, affect experimental precision, as the CFX96Touch quantitative real time PCR Instrument of BIO-RAD company;
2. the light path design of some fluorescent quantitative PCR detector is very complicated, this adds increased volume and the cost of product, reduces reliability, as the TOpticalThermocycler quantitative real time PCR Instrument of Biometra company;
3. some fluorescent quantitative PCR detector is by increasing rotating exciting light optical filter colour wheel and utilizing emitted light optical filter colour wheel in the optical path, reach the object that multi-pass detects, the existence of these mechanical hook-ups adds lower power consumption reliability, as the LightCycler480 quantitative real time PCR Instrument of Roche company;
4. in addition, most fluorescent quantitative PCR detectors are in the market the medium-and-large-sized machines for test in laboratory, and expensive, portability is poor, the sample prepared must be taken in laboratory and detect by user, and this mode have impact on the quick diagnosis of some sudden illness.
In view of this, the present inventor studies this, and develop a kind of Portable fluorescence quantitative PCR detector specially, this case produces thus.
Summary of the invention
The object of this invention is to provide and a kind ofly carry out that multispectral passage Multi-example detects simultaneously, mechanical mobile device, to detect flexibly and the low Portable fluorescence quantitative PCR detector of cost.
To achieve these goals, solution of the present invention is:
A kind of Portable fluorescence quantitative PCR detector, comprise optical excitation unit, optical detection unit and temperature control unit, wherein said optical excitation unit comprises several light sources, be arranged on the exciting light optical filter on each light source light direction, and the completely reflecting mirror be inclined to set with the exciting light that light source penetrates; Described temperature control unit comprises reaction tank for placing PCR test tube, be arranged on the heat lid at PCR test tube top, be arranged on heating/refrigerating sheet bottom PCR test tube, be arranged on thermal component below heating/refrigerating sheet, and for regulating heat lid and heating/refrigerating sheet temperature temperature-control circuit just; Described optical detection unit comprises several light splitting pieces, optical filter, imaging sensor, and for the testing circuit of control both optical imaging.The light that light source sends becomes the exciting light of specific wavelength after exciting light optical filter, inject in PCR test tube after completely reflecting mirror reflection again, the fluorescent material that exciting light irradiates in reagent produces utilizing emitted light, a part utilizing emitted light enter optical detection unit by the lateral opening hole of reaction tank, eventually pass the light splitting of optical detection unit, filter after imaging on corresponding imaging sensor.
As preferably, above-mentioned light source comprises reflector and is arranged on the monochromatic LED at reflector center, light source adopts monochromatic LED, have compared with traditional halogen tungsten lamp and generate heat less, wavelength utilization factor is high, and volume is little, the advantage that the life-span is long, and LED light source energy obtains utilizing the most efficiently to adopt reflector to ensure, also can ensure that exciting light keeps collimation evenly simultaneously.
For ensureing the homogeneity of light source, as preferably, the light path between the light source of described fluorescent quantitative PCR detector and PCR test tube being provided with array lens group further, the brightness of whole bundle light at each position can being made all relatively consistent by array lens group.
As preferably, above-mentioned exciting light optical filter adopts centre wavelength at the bandpass filter of 400 ~ 650nm.
As preferably, described completely reflecting mirror and horizontal direction are inclined to set, and make exciting light after total reflection, vertically inject in PCR test tube, ensure the exciting light after total reflection and utilizing emitted light at an angle, thus reduction utilizing emitted light ground unrest.In order to reduce the interference between exciting light and utilizing emitted light, exciting light light path and utilizing emitted light light path preferably angled, and the interference at right angle is minimum, uses the light microscopic that is all-trans can the most effectively utilize space under the prerequisite ensureing utilizing emitted light light path best angle, accomplish complete machine miniaturization.
As preferably, described optical detection unit is provided with multiple detection light path, each detection light path has the light splitting piece of corresponding wavelength successively, optical filter, the composition such as lens and imaging sensor, after entering optical detection unit from the utilizing emitted light of multiple PCR test tube injection, first carry out light splitting through the first light splitting piece, the light beam of the first bundle specific wavelength separated injects the first corresponding optical filter and the first lens, last imaging on a first imaging sensor, remaining utilizing emitted light carries out secondary light splitting through the second light splitting piece again, the light separating the second bundle specific wavelength injects the second corresponding optical filter and the second lens, last imaging on the second imaging sensor, light splitting successively as stated above, until the in the end imaging on an imaging sensor of last road light beam.
As preferably, the angle between the utilizing emitted light of each PCR test tube injection and the primary optical axis of optical detection unit is α, wherein 0≤α≤12 °.Because the existence of α, the spectrum of light splitting piece and optical filter can be caused to depart to shortwave, so just likely make exciting light through interfering with test result, for avoiding this puzzlement, needing α to control within certain angle.
As preferably, above-mentioned imaging sensor adopts highly sensitive cmos imaging sensor or ccd imaging sensor, such as the EXiBlueCCDImager of the ULS24CMOSImager of anitoa company, QImaging company.
As preferably, the reaction tank of described temperature control unit comprises base plate and is arranged on several cuvette grooves on base plate, the sidewall of described cuvette groove just to optical detection unit direction is equipped with a light hole, for the invisible spectro radiative injection of PCR, the lower limb of light hole with to flush bottom cuvette groove or higher than 0-2mm bottom cuvette groove.The internal diameter of described cuvette groove and the external diameter of PCR test tube match, and make PCR test tube outer wall be close to cuvette groove inwall, can increase contact area like this and be conducive to heat transfer.
As preferably, described reaction tank adopts silver, copper, aluminium material, and reaction tank base plate lower surface is smooth flat, is conducive to conducting heat because polish smooth.Base plate surrounding is coated with heat-barrier material, can reduce the thermal losses that cross-ventilation causes.
As preferably, described heating/refrigerating sheet is Peltier, is arranged on the lower surface of reaction tank, reaches the object of heating and refrigeration by changing Peltier direction of current.
As preferably, the thermal component of described temperature control unit comprises heating radiator, fan etc., and described thermal component is arranged on the below of heating/refrigerating sheet, for dispersing the heat that Peltier produces.The form of heating radiator is varied, and key will make air channel unimpeded, and heat can discharge instrument in time.
As preferably, the upper and lower surface of described heating/refrigerating sheet is provided with thermally-conductive interface packing material further, makes heat in Peltier and heating radiator, good communication between Peltier and reaction tank base plate.Thermally-conductive interface packing material will select the material that thermal conductivity is high, and shape is as far as possible thin, and there is not air centre, recommends but is not limited to graphite guide hotting mask, heat-conducting silicone grease.
As preferably, described heat lid comprises aluminium sheet and is pasted onto the flexible electrical heating film of aluminium sheet upper surface, and in testing process, heat lid is pressed on test tube cap and keeps certain pressure.The hot temperature covered is higher than the temperature of reaction tank.At 95 degrees centigrade during the high temperature of General reactions pond, heat lid then keeps about 105 DEG C, so both can prevent water vapor from condensing on test tube cap, again can to test tube cap pressurization prevent test tube interior pressure too senior general's lid pop, light hole is left, injecting for exciting light in the position of the corresponding test tube of heat lid.
As preferably, said temperature control circuit comprises the hot lid temperature sensor being separately positioned on heat and covering, reaction tank temperature sensor, microprocessor and driving circuit, wherein, hot lid temperature sensor is connected with the input end of microprocessor respectively with reaction tank temperature sensor, driving circuit one end is connected with the output terminal of microprocessor, the other end connects heat lid and heating/refrigerating sheet respectively, temperature sensor and reaction tank temperature sensor Real-time Collection heat lid and reaction tank on temperature, and feed back to microprocessor, microprocessor is after data processing, instruction is sent to driving circuit, the heating intensity of heat lid and heating/refrigerating sheet is controlled by driving circuit, thus make PCR test tube control in suitable range of reaction temperature, the mode of PWM is regulated to carry out to the driving available pulse widths of hot lid well heater and heating/refrigerating sheet, PWM frequency should higher than best more than the 100Hz of 50Hz..
As preferably, the testing circuit of above-mentioned optical detection unit comprises image acquisition and process microprocessor and LED drive circuit, the input end of microprocessor is connected with imaging sensor, and output terminal is connected with LED drive circuit, and the output terminal of described LED drive circuit is connected with each monochromatic LED respectively.After a temperature cycles, optical detecting module is started working, and microprocessor sends to LED drive circuit the LED that multiple wave band is lighted in instruction, sends exciting light.Microprocessor starts the image gathering imaging sensor on each passage simultaneously, then image is processed to the fluorescence intensity obtaining each test tube.
Fluorescent quantitative PCR detector of the present invention, with the LED of low power consumption and small volume as excitation source, be irradiated on all reaction tanks simultaneously, and with highly sensitive CMOS or ccd imaging sensor as detector, the imaging while that all reaction tanks being carried out.Use optical filter and light splitting piece by the imaging simultaneously in several light path of the light of different wave length simultaneously, thus multiple different fluorescent material can be detected in a sample, can detect multiple sample simultaneously.No longer need huge mechanical scanner to each sample one by one inspection, simplify system, avoid the inaccuracy because sample optical check time difference causes, be specially adapted to the small-sized solution of detection of nucleic acids fast.In addition, compared with prior art, also tool has the following advantages above-mentioned fluorescent quantitative PCR detector:
(1) PCR detector mechanical gearing of the present invention, avoids the mechanical fault problem of later stage use procedure;
(2) by temperature control unit, temperature is accurately controlled in range of reaction temperature, make testing result more accurate;
(3) all samples detects simultaneously, avoids the error brought because of the mistiming scanned, and improves the repeatability detected;
(4) compact, can carry movement, convenient detects on the spot, decreases sample and transports the time cost that brings and sample degradation and to go bad risk;
(5) greatly reduce the price threshold of real-time fluorescence PCR instrument, improve the popularity rate of round pcr.
Below in conjunction with drawings and the specific embodiments, the present invention is described in further detail.
Accompanying drawing explanation
Fig. 1 is the fluorescent quantitative PCR detector inner structure schematic diagram of the present embodiment;
Fig. 2 is the fluorescent quantitative PCR detector sectional view of the present embodiment;
Fig. 3 is the inner light path schematic diagram of fluorescent quantitative PCR detector optical detection unit of the present embodiment;
Fig. 4 is the fluorescent quantitative PCR detector temperature-control circuit driving circuit principle figure of the present embodiment;
Fig. 5 is the fluorescent quantitative PCR detector testing circuit microprocessor schematic diagram of the present embodiment.
Embodiment
As shown in Figure 1, a kind of Portable fluorescence quantitative PCR detector, comprise optical excitation unit 1, optical detection unit 3 and temperature control unit 2, wherein said optical excitation unit 1 comprises several light sources 11, be arranged on the exciting light optical filter 12 on each light source 11 light direction, and the completely reflecting mirror 13 be inclined to set with the exciting light that light source penetrates; Described temperature control unit 2 comprises reaction tank 21 for placing PCR test tube 4, be arranged on the heat lid 22 at PCR test tube 4 top, be arranged on heating/refrigerating sheet 23 bottom PCR test tube 4, be arranged on thermal component 24 below heating/refrigerating sheet 23, and for regulating heat lid and heating/refrigerating sheet 23 temperature temperature-control circuit just; Described optical detection unit 3 comprises several light splitting pieces, optical filter, imaging sensor, and for the testing circuit of control both optical imaging.The light that light source 11 sends becomes the exciting light of specific wavelength after exciting light optical filter 12, inject in PCR test tube 4 after completely reflecting mirror 13 reflects again, the fluorescent material that exciting light irradiates in reagent produces utilizing emitted light, a part utilizing emitted light enter optical detection unit 3 by the lateral opening hole 213 of reaction tank 21, eventually pass the light splitting of optical detection unit 3, filter after imaging on corresponding imaging sensor.
In the present embodiment, as shown in Figure 2, described Portable fluorescence quantitative PCR detector comprises 3 sense channels, and therefore described light source 11 comprises 3 reflectors 111 and is arranged on the monochromatic LED 112 at each reflector 111 center, and light source 11 adopts monochromatic LED 112, have compared with traditional halogen tungsten lamp and generate heat less, wavelength utilization factor is high, and volume is little, the advantage that the life-span is long, and LED light source energy obtains utilizing the most efficiently to adopt reflector 111 to ensure, also can ensure that exciting light keeps collimation evenly simultaneously.For ensureing the homogeneity of light source 11, the light path between light source 11 and PCR test tube 4 can install array lens group further, the brightness of whole bundle light at each position can be made all relatively consistent by array lens group.
In order to reduce the interference between exciting light and utilizing emitted light, exciting light light path and utilizing emitted light light path preferably angled, and the interference at right angle is minimum, therefore, completely reflecting mirror 13 described in the present embodiment is 45 ° with horizontal direction and is obliquely installed, horizontal direction is made to penetrate next exciting light after total reflection, vertically inject in PCR test tube 4, fluorescent material produces utilizing emitted light level injection again, ensure total reflection after exciting light and utilizing emitted light rectangular, use the light microscopic 13 that is all-trans can the most effectively utilize space under the prerequisite ensureing utilizing emitted light light path best angle, accomplish complete machine miniaturization.
In the present embodiment, described exciting light optical filter 12 adopts the bandpass filter of 3 different centre wavelengths, specifically as shown in table 1.
The exciting light of table 1 three passages and utilizing emitted light centre wavelength
| Exciting light centre wavelength (nm) | Utilizing emitted light centre wavelength (nm) | Detectable fluorescent material citing |
| 470 | 510 | FAM，SYBR Green I,Fluorescein,EvaGreen,Alexa Fluor 488 |
| 530 | 550 | JOE,VIC,HEX,TET,MAX,CAL Fluor,Gold 540,Yakima Yellow |
| 570 | 610 | ROX,CAL Fluor Red 610,Cy3.5,Texas Red, lexa Fluor 568 |
The reaction tank 21 of described temperature control unit 2 comprises base plate 211 and 3 cuvette grooves 212 be arranged on base plate 211, the sidewall of described cuvette groove 212 just to optical detection unit 3 direction is equipped with a light hole 213, for the radiative injection in PCR test tube 4, the lower limb of light hole 213 with to flush bottom cuvette groove or higher than 0-2mm bottom cuvette groove.The internal diameter of described cuvette groove 212 and the external diameter of PCR test tube 4 match, and make PCR test tube 4 outer wall be close to cuvette groove inwall, can increase contact area like this and be conducive to heat transfer.Described reaction tank 21 adopts silver, copper, aluminium material, and the present embodiment selects fine aluminium, and general fine aluminium is better than copper, this is because the aluminium of same volume and copper ratio of specific heat 2.376:3471, the heat namely raising identical temperature aluminium needs is less.Reaction tank 21 is as far as possible little in content with funtion requirement situation, improves Cooling rate to reduce thermal capacitance.The metal reaction pond design that the present embodiment provides is through overheated emulation and experiment, and the temperature homogeneity between each test tube is good, and the temperature difference is less than 0.3 DEG C.Reaction tank base plate 211 lower surface is smooth flat, is conducive to heat transfer because polish smooth.Base plate 211 surrounding is coated with heat-barrier material, can reduce the thermal losses that cross-ventilation causes.
In the present embodiment, described heating/refrigerating sheet 23 is Peltier, is arranged on the lower surface of reaction tank 21, and reach the object of heating and refrigeration by changing Peltier direction of current, its physical principle is peltier effect.The function of heating and refrigeration is reached, the device power > load thermal capacitance * Cooling rate selected by changing direction of current.The upper and lower surface of described heating/refrigerating sheet 23 is provided with thermally-conductive interface packing material further, makes heat in Peltier and heating radiator, good communication between Peltier and reaction tank base plate.Thermally-conductive interface packing material will select the material that thermal conductivity is high, and shape is as far as possible thin, and there is not air centre, recommends but is not limited to graphite guide hotting mask, heat-conducting silicone grease.
The thermal component 24 of described temperature control unit comprises by heating radiator, fan etc., and described thermal component 24 is arranged on the below of Peltier, for dispersing the heat that Peltier produces.The form of heating radiator is varied, and key will make air channel unimpeded, and heat can discharge instrument in time.
Described heat lid 22 comprises aluminium sheet 222 and is pasted onto the flexible electrical heating film 221 of aluminium sheet 222 upper surface, and in testing process, heat lid 22 is pressed on test tube cap and keeps certain pressure.The temperature of heat lid 22 is higher than the temperature of reaction tank.At 95 degrees centigrade during 21 high temperature of General reactions pond, 22, heat lid keeps about 105 DEG C, so both can prevent water vapor from condensing on test tube cap, again can to test tube cap pressurization prevent test tube interior pressure too senior general's lid pop, light hole is left, injecting for exciting light in the position of heat lid 22 corresponding test tubes 4.
As shown in Figure 3, described optical detection unit 3 is provided with 3 and detects light path, each detection light path has the light splitting piece of corresponding wavelength successively, optical filter, lens and imaging sensor composition, after entering optical detection unit 3 from the utilizing emitted light of multiple PCR test tube 4 injection, first carry out light splitting through the first light splitting piece 31, the light beam of the first bundle specific wavelength (510nm) separated injects the first corresponding optical filter 33 and the first lens 36, last imaging on the first imaging sensor 39, remaining utilizing emitted light carries out secondary light splitting through the second light splitting piece 32 again, the light separating the second bundle specific wavelength (550nm) injects the second corresponding optical filter 34 and the second lens 37, last imaging on the second imaging sensor 40, the light of remaining three beams specific wavelength (610nm) injects the 3rd corresponding optical filter 35 and the 3rd lens 38, last imaging on the 3rd imaging sensor 41.
Angle between the utilizing emitted light of each PCR test tube 4 injection and the primary optical axis of optical detection unit 3 is α, wherein 0≤α≤12 °.Because the existence of α, the spectrum of the light splitting piece in optical detection unit 3 and optical filter can be caused to depart to shortwave, so just likely make exciting light through interfering with test result, for avoiding this puzzlement, needing α to control within certain angle.
Above-mentioned first imaging sensor 39, second imaging sensor 40 and the 3rd imaging sensor 41 adopt highly sensitive cmos imaging sensor, in the present embodiment, the ULS24CMOSImager of concrete employing anitoa company, above-mentioned imaging sensor also can adopt ccd imaging sensor, such as the EXiBlueCCDImager of QImaging company.
As preferably, said temperature control circuit comprises the hot lid temperature sensor 223 be arranged on heat lid 22, PT-100 platinum thermal resistance sensor can be adopted, reaction tank temperature sensor 214, TSIC201WST high-precision sensor can be adopted, microprocessor (adopting STM32F103ZE model) and driving circuit, as shown in Figure 4, wherein, hot lid temperature sensor 223 is connected with the input end of microprocessor respectively with reaction tank temperature sensor 214, driving circuit one end is connected with the output terminal of microprocessor, the other end connects heat lid 22 and heating/refrigerating sheet 23 respectively, hot lid temperature sensor 223 and reaction tank temperature sensor 214 Real-time Collection heat cover the temperature on 22 and reaction tank 21, and feed back to microprocessor, microprocessor is after data processing, instruction is sent to driving circuit, the heating intensity of heat lid 22 and heating/refrigerating sheet 23 is controlled by driving circuit, thus make PCR test tube 4 control in suitable range of reaction temperature, the mode of PWM is regulated to carry out to the driving available pulse widths of hot lid well heater and heating/refrigerating sheet, PWM frequency should higher than best more than the 100Hz of 50Hz..
The testing circuit of above-mentioned optical detection unit 3 comprises image acquisition and process microprocessor and LED drive circuit, image acquisition is connected with each imaging sensor with the input end of process microprocessor, output terminal is connected with LED drive circuit, and the output terminal of described LED drive circuit is connected with each monochromatic LED respectively.After a temperature cycles, optical detecting module is started working, and image acquisition sends to LED drive circuit the LED that multiple wave band is lighted in instruction with process microprocessor, sends exciting light.Microprocessor starts the image gathering imaging sensor on each passage simultaneously, then image is processed to the fluorescence intensity obtaining each test tube.The microprocessor of temperature-control circuit and the microprocessor of imaging moiety testing circuit can be same components and parts, also the microprocessor of two same models or different model can be adopted respectively, if both are not same microprocessors, communication coordination need be carried out by existing communication protocol, both can Direct Communication, or through upper machine communication.In the present embodiment, the microprocessor of imaging moiety testing circuit adopts the microprocessor chip of same STM32F103ZE model, as shown in Figure 5.In addition, fluorescent quantitative PCR detector described in the present embodiment by arranging the communication module (as bluetooth module etc.) be connected with process microprocessor with image acquisition, can also realize the communication of fluorescent quantitative PCR detector and other equipment (mobile phone, host computer etc.).
The present embodiment adopts three-passage design, can detect three kinds of fluorescent materials in a sample simultaneously.Principle of work is as follows:
470nm monochromatic LED is opened, and reflexed on the reagent in test tube 4 by total reflective mirror 13 after exciting light optical filter 12, the sample with FAM fluorescent material is excited, and sends the utilizing emitted light of about 510nm.Utilizing emitted light is reflected by the first light splitting piece 31 (the long reduction of fractions to a common denominator mating plate of 530nm) after entering optical detection unit 3, imaging on the first imaging sensor 39 after utilizing emitted light bandpass filter, check that shape and the brightness of picture can judge which sample contains FAM fluorescent material, and concentration.
530nm monochromatic LED is opened, and reflexed on the reagent in test tube 4 by total reflective mirror 13 after exciting light optical filter 12, the sample with JOE fluorescent material is excited, and sends the utilizing emitted light of about 550nm.Utilizing emitted light passes the first light splitting piece 31 after entering optical detection unit 3, then by the long reduction of fractions to a common denominator mating plate of the second light splitting piece 32(585nm) reflection, imaging on the second imaging sensor 40 after utilizing emitted light bandpass filter, check that shape and the brightness of picture can judge which sample contains JOE fluorescent material, and concentration.
570nm monochromatic LED is opened, and reflexed on invisible spectro reagent by total reflective mirror 13 after exciting light optical filter 12, the sample with ROX fluorescent material is excited, and sends the utilizing emitted light of about 610nm.Utilizing emitted light passes the first light splitting piece 31 and the second light splitting piece 32 after entering optical detection unit 3, imaging on the 3rd imaging sensor 41 after utilizing emitted light bandpass filter, check that shape and the brightness of picture can judge which sample contains ROX fluorescent material, and concentration.
The present embodiment is illustrated with triple channel, but is not limited to three passages, can have 2 passages, 4 passages, 5 passages etc., with FAM, JOE, ROX tri-kinds of fluorescent material citings, but it is several to be not limited to this, if the SYBRGreenI that FAM as shown in table 1 changes identical service band into is also passable.
Fluorescent quantitative PCR detector described in the present embodiment, with the LED of low power consumption and small volume as excitation source, be irradiated on all reaction tanks simultaneously, and with high sensitivity CMOS or ccd imaging sensor as detector, the imaging while that all reaction tanks being carried out.Use optical filter and light splitting piece by the imaging at one time respectively in several light path of the light of different wave length simultaneously, thus multiple different fluorescent material can be detected in a sample, and can detect multiple sample simultaneously.No longer need huge mechanical scanner to each sample one by one inspection, simplify system, avoid the inaccuracy because sample optical check time difference causes, be specially adapted to the small-sized solution of detection of nucleic acids fast.
Above-described embodiment and graphic and non-limiting product form of the present invention and style, any person of an ordinary skill in the technical field, to its suitable change done or modification, all should be considered as not departing from patent category of the present invention.
Claims (16)
1. a Portable fluorescence quantitative PCR detector, it is characterized in that: comprise optical excitation unit, optical detection unit and temperature control unit, wherein said optical excitation unit comprises several light sources, be arranged on the exciting light optical filter on each light source light direction, and the completely reflecting mirror be inclined to set with the exciting light that light source penetrates; Described temperature control unit comprises reaction tank for placing PCR test tube, be arranged on the heat lid at PCR test tube top, be arranged on heating/refrigerating sheet bottom PCR test tube, be arranged on thermal component below heating/refrigerating sheet, and for regulating heat lid and heating/refrigerating sheet temperature temperature-control circuit just; Described optical detection unit comprises several light splitting pieces, optical filter, imaging sensor, and for the testing circuit etc. of control both optical imaging; The light that light source sends becomes the exciting light of specific wavelength after exciting light optical filter, inject in PCR test tube after completely reflecting mirror reflection again, the fluorescent material that exciting light irradiates in reagent produces utilizing emitted light, a part utilizing emitted light enter optical detection unit by the lateral opening hole of reaction tank, eventually pass the light splitting of optical detection unit, filter after imaging on corresponding imaging sensor.
2. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, is characterized in that: described light source comprises reflector and is arranged on the monochromatic LED at reflector center.
3. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, it is characterized in that: the light path between the light source of described fluorescent quantitative PCR detector and PCR test tube is provided with array lens group further, make the brightness of whole bundle light at each position all relatively consistent by array lens group.
4. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, is characterized in that: described exciting light optical filter adopts centre wavelength at the bandpass filter of 400 ~ 650nm.
5. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, is characterized in that: described completely reflecting mirror and horizontal direction are inclined to set, and makes exciting light after total reflection, vertically injects in PCR test tube.
6. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, it is characterized in that: described optical detection unit is provided with multiple detection light path, each detection light path mainly contains the light splitting piece of corresponding wavelength successively, optical filter, lens and imaging sensor composition, after entering optical detection unit from the utilizing emitted light of multiple PCR test tube injection, first carry out light splitting through the first light splitting piece, the light beam of the first bundle specific wavelength separated injects the first corresponding optical filter and the first lens, last imaging on a first imaging sensor, remaining utilizing emitted light carries out secondary light splitting through the second light splitting piece again, the light separating the second bundle specific wavelength injects the second corresponding optical filter and the second lens, last imaging on the second imaging sensor, light splitting successively as stated above, until the in the end imaging on an imaging sensor of last road light beam.
7. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 6, it is characterized in that: the angle between the utilizing emitted light of each PCR test tube injection and the primary optical axis of optical detection unit is α, wherein 0≤α≤12 °, to ensure that the operation wavelength that light splitting piece and optical filter cause because incident angle departs from departs from less.
8. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, is characterized in that: described imaging sensor adopts cmos imaging sensor or ccd imaging sensor.
9. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, it is characterized in that: the reaction tank of described temperature control unit comprises base plate and is arranged on several cuvette grooves on base plate, the sidewall of described cuvette groove just to optical detection unit direction is equipped with a light hole, for the invisible spectro radiative injection of PCR, the lower limb of light hole with to flush bottom cuvette groove or higher than 0-2mm bottom cuvette groove.
10. a kind of Portable fluorescence quantitative PCR detector as claimed in claim 1, is characterized in that: described reaction tank adopts silver, copper, aluminium material, and reaction tank base plate lower surface is smooth flat.
11. a kind of Portable fluorescence quantitative PCR detectors as claimed in claim 1, is characterized in that: described heating/refrigerating sheet is Peltier, is arranged on the lower surface of reaction tank.
12. a kind of Portable fluorescence quantitative PCR detectors as claimed in claim 1, it is characterized in that: the thermal component of described temperature control unit comprises heating radiator, fan, described thermal component is arranged on the below of heating/refrigerating sheet, for dispersing the heat that Peltier produces.
13. a kind of Portable fluorescence quantitative PCR detectors as claimed in claim 11, is characterized in that: the upper and lower surface of described heating/refrigerating sheet is provided with thermally-conductive interface packing material further.
14. a kind of Portable fluorescence quantitative PCR detectors as claimed in claim 1, is characterized in that: described heat lid comprises aluminium sheet and is pasted onto the flexible electrical heating film of aluminium sheet upper surface.
15. a kind of Portable fluorescence quantitative PCR detectors as claimed in claim 1, it is characterized in that: said temperature control circuit comprises the hot lid temperature sensor, reaction tank temperature sensor, microprocessor and the driving circuit that are separately positioned on heat and cover, wherein, temperature sensor is connected with the input end of microprocessor respectively with reaction tank temperature sensor, driving circuit one end is connected with the output terminal of microprocessor, and the other end connects heat lid and heating/refrigerating sheet respectively; Hot lid temperature sensor and reaction tank temperature sensor Real-time Collection heat lid and reaction tank on temperature, and feed back to microprocessor, microprocessor is after data processing, instruction is sent to driving circuit, controlled the heating intensity of heat lid and heating/refrigerating sheet by driving circuit, thus PCR test tube is controlled in suitable range of reaction temperature.
16. a kind of Portable fluorescence quantitative PCR detectors as claimed in claim 1, it is characterized in that: the testing circuit of above-mentioned optical detection unit comprises image acquisition and process microprocessor and LED drive circuit, the input end of microprocessor is connected with imaging sensor, output terminal is connected with LED drive circuit, the output terminal of described LED drive circuit is connected with each monochromatic LED respectively, microprocessor sends instruction to driving circuit, light LED and produce exciting light, the utilizing emitted light signal of microcontroller acquires imaging sensor collection simultaneously.
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Cited By (31)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105400691A (en) * | 2015-12-11 | 2016-03-16 | 舟山医院 | A multifunctional PCR instrument |
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| CN106929388A (en) * | 2015-12-31 | 2017-07-07 | 苏州百源基因技术有限公司 | A kind of real-time fluorescence quantitative PCR instrument |
| CN105524826A (en) * | 2016-01-12 | 2016-04-27 | 天津喜诺生物医药有限公司 | Detection device of QPCR multichannel mobile light source |
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| CN106680474A (en) * | 2017-02-04 | 2017-05-17 | 上海为然环保科技有限公司 | Instrument capable of quickly detecting escherichia coli on household tableware |
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| CN108181284A (en) * | 2018-01-19 | 2018-06-19 | 杭州安誉科技有限公司 | A kind of data acquisition device of real-time fluorescence quantitative PCR instrument |
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| CN109060742B (en) * | 2018-08-06 | 2021-10-26 | 张家林 | Portable thermal cycle fluorescence detector |
| CN110327992B (en) * | 2019-02-03 | 2020-11-06 | 北京理工大学 | Heating device and portable QPCR device using same |
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| US11767552B2 (en) | 2019-03-21 | 2023-09-26 | Biomeme, Inc. | Multi-function analytic devices |
| CN110218647A (en) * | 2019-06-27 | 2019-09-10 | 杭州安塔生物科技有限公司 | A kind of PCR reaction unit |
| CN110320190A (en) * | 2019-07-04 | 2019-10-11 | 欧蒙医学诊断（中国）有限公司 | A kind of fluorescence detector |
| CN111013688B (en) * | 2019-12-06 | 2022-05-17 | 深圳市刚竹医疗科技有限公司 | qPCR module and modularized qPCR device |
| CN111013688A (en) * | 2019-12-06 | 2020-04-17 | 深圳市刚竹医疗科技有限公司 | qPCR module and modularized qPCR device |
| CN111592982A (en) * | 2020-05-24 | 2020-08-28 | 杭州天微基因科技有限公司 | Novel portable nucleic acid amplification analyzer |
| CN113832020A (en) * | 2020-06-24 | 2021-12-24 | 傅宗民 | Optical module, thermal cycle module and PCR device suitable for PCR device |
| CN111751341A (en) * | 2020-06-24 | 2020-10-09 | 新乡市食品药品检验所 | Traditional Chinese medicinal material component fluorescence analysis detector and analysis detection method |
| US11892461B1 (en) | 2020-09-18 | 2024-02-06 | Biomeme, Inc. | Portable devices and methods for analyzing samples |
| CN114381361A (en) * | 2020-10-19 | 2022-04-22 | 台达电子国际（新加坡）私人有限公司 | Light-activated devices and methods of controlling same |
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| CN112709973A (en) * | 2020-12-31 | 2021-04-27 | 深圳市必拓电子股份有限公司 | Method for generating composite light by exciting fluorescence with laser and light path structure thereof |
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