CN103207169A - Microfluidic time-resolved fluorescence immunoassay device and application thereof - Google Patents
Microfluidic time-resolved fluorescence immunoassay device and application thereof Download PDFInfo
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Abstract
The invention provides a microfluidic time-resolved fluorescence immunoassay device. The device comprises a sample injection unit (101), a microfluidic control unit (103), a detection unit (104) and a microfluidic chip (102) adopting a membrane pump drive mode, and the microfluidic chip (102) is positioned in a microfluidic chip carrier on an analysis workbench of the microfluidic control unit (103). The microfluidic time-resolved fluorescence immunoassay device has the advantages that by adopting the microfluidic technology, a detection system is simplified; and a microplate oscillator and a microplate washer are omitted, a microfluidic control system is simple in structure, optimized, free of mechanical movements and structures of cleaning, vibration, transmission and the like, and automation is achieved conveniently.
Description
Technical field
The invention belongs to field of immunology, particularly time-resolved fluorescence analytical equipment and application thereof.
Background technology
The time-resolved fluorescence analytical technology is to have the lanthanide series of unique fluorescent characteristic and sequestrant thereof as tracer, a kind of novel on-radiation trace analysis of foundation.TRFIA methodological study and clinical practice development are rapidly, become a new milestone of labelled immune analysis development, highly sensitive, advantages such as standard curve range is wide, easy and simple to handle, "dead" pollution, multiple labeling that the TRFIA technology has, be widely used in the clinical trial diagnosis of tumour, infectious disease, endocrine system disease, autoimmune disease, genetic disease, become one of analysis means commonly used in biomedical research and the clinical ultramicron biochemical investigation.
The TRFIA(time resolved fluoro-immunoassay) utilized 3 valency rare earth ions and chelate thereof with unique fluorescent characteristic to replace fluorescent material, enzyme, isotope, chemiluminescent substance for tracer, labelled antibody, antigen, hormone, polypeptide, protein, nucleic acid probe and biological cell, after question response system (as: antigen-antibody reaction, nucleic acid probe hybridization, the reaction of biotin Avidin and target cell pairing effect cell kill and wound reaction etc.) takes place, with the fluorescence intensity in the TRFIA detector assaying reaction product.According to the ratio of product fluorescence intensity and relative intensity of fluorescence, judge the concentration of analyte in the reaction system, thereby reach quantitative test.
In the time-resolved fluoroimmunoassay detection method of prior art, the general detection finished at 96 microwell plates, a complete optimization system preparation should comprise an automatic pipettor, a microwell plate oscillator, a microwell plate is washed the plate machine, a couveuse, a time resolved fluoro-immunoassay instrument; Or the full-automatic time resolved fluoro-immunoassay instrument of an integrated above-mentioned functions.The detection step is generally: 1) → add sample, add sample-A 50 μ l; 2) → adding dilution-B 100 μ l; 3) → and vibration, antigen-antibody (bag quilt) reaction bonded, about 60min; 4) → and wash plate, it is empty to add cleaning fluid-suction, and it is empty to add cleaning fluid-suction again, 2～6 times; Or manual the cleaning, buckle is cleaned; 5) → adding europium marking fluid-C 150 μ l; 6) → and vibration, " tape label antibody "+" antigen-antibody (bag quilt) reaction bonded body forms sandwich ", about 30min; 7) → wash plate, with step 4; 8) → add enhancing liquid-D 150 μ l that dissociate; 9) → and vibration, the europium that dissociates, about 5min; 10) → detect.All testing process is carried out 4 step application of samples, and (96*4=376 time altogether) washes plate 2 times, vibrates about 150～180min of time spent 3 times.And carry out the basic support equipment of semi-automatic detection be: a microwell plate autopipette, microwell plate oscillator, microwell plate are washed the plate machine.Prior art is used semi-automatic checkout equipment, and the time that needs is longer, more take manpower.
Full-automatic time resolved fluoro-immunoassay instrument refers to that time-resolved fluoroimmunoassay is tested (comprise application of sample, hatch, wash plate, add operations such as strengthening liquid) and the detection of fluorescence is successively all finished on whole instrument, need artificial participation hardly; Though saved the loaded down with trivial details and inaccuracy of artificial application of sample, especially when the detection of great amount of samples, but existed following major defect: apparatus structure complexity, instrument costliness; Instrument uses, the maintenance requirement height, and comprehensive use cost height can't be popularized use.
And semi-automatic detection system configuration needs 4 kinds of equipment: one " microwell plate autopipette ", one " microwell plate oscillator ", one " microwell plate rinsing maching ", a time resolved fluoro-immunoassay instrument.It also has following defective: the detection system complexity, use 4 kinds of equipment; Instrument uses, the maintenance requirement height, and comprehensive use cost height can't be popularized use.
Summary of the invention
In order to address the above problem, the objective of the invention is to adopt time-resolved fluorescence analytical technology and film to move the micro-fluidic chip technology in the application of biological chemistry and molecular biology application, a kind of combination of using the moving micro-fluidic chip technology of film and time-resolved fluoroimmunoassay detection technique is provided especially, realizes full-automatic micro-fluidic time-resolved fluoroimmunoassay detection method.
In order to realize the object of the invention, concrete technical scheme is:
A kind of micro-fluidic time resolved fluoro-immunoassay device, comprise application of sample unit, micro-fluidic control module, detecting unit, it also comprises the micro-fluidic chip that adopts the membrane pump type of drive, and described micro-fluidic chip is arranged in the micro-fluidic chip seat on the micro-fluidic control module analytical work platform.
Wherein, described micro-fluidic chip comprises: sample pool, dilution liquid pool, mark liquid pool, the liquid pool that dissociates, cleaning liquid pool, waste liquid pool;
Be provided with sample valve and sample through hole in the wherein said sample pool; Be provided with dilution valve and dilution through hole in the dilution liquid pool; Be provided with marking fluid valve and marking fluid through hole in the described mark liquid pool, be provided with the dissociation solution valve in the described liquid pool that dissociates and the fluid through-hole that dissociates, described cleaning liquid pool is provided with cleaning fluid valve and cleaning fluid through hole; Described waste liquid pool is provided with waist valve and waste liquid through hole;
Main valve is arranged at the centre of each valve, and each valve links to each other with main valve respectively by passage.
Wherein, described sample valve, main valve and dilution liquid pool valve and passage constitute sample pool-dilution liquid pool two-way pump sample dilution pumps; Described sample valve, main valve and waist valve and passage constitute sample pool-waste liquid pool one-way pump sample waste drains pump; Described sample valve, main valve and cleaning fluid valve and passage constitute sample pool-cleaning liquid pool one-way pump sample scavenging pump; Described sample valve, main valve and marking fluid valve and passage constitute sample pool-mark liquid pool two-way pump sample labeling pump; Described sample valve, main valve and dissociation solution valve and passage constitute the sample pool-liquid pool two-way pump sample solution that dissociates from pump.
Wherein, described sample pool inwall is coated with antibody or antigen.
Wherein, described micro-fluidic chip is that light tight polymeric material is made.Be preferably polystyrene (PS) or acrylonitrile-butadiene-styrene (ABS) plastics (ABS).
Wherein, described application of sample unit includes three-dimensional motion mechanism, at three-dimensional motion mechanism feed head and sample-adding pump is arranged; Three-dimensional motion mechanism moves in analytical work platform scope; Described application of sample unit also includes reagent cup, and described reagent cup comprises the dilution cup, the marking fluid cup, and the dissociation solution cup, the cleaning fluid cup all is placed on the device analysis work top.
Wherein, described micro-fluidic control module comprises single-chip microcomputer, control circuit system, and control is finished equipment and operated the communication that reaches with external unit; Chip carrier and chip are complementary, and are furnished with drives structure in the position corresponding with the micro-fluidic chip membrane pump, are used for the driving to the micro-fluidic chip membrane pump.
The membrane pump mechanical work principle as shown in Figure 9.When departing from its position originally, the barrier film stress deformation just formed the diaphragm seal chamber.This film chamber is linked up two passages, makes that liquid is inhaled into the film chamber in the passage.By six continuous step cycle work, realize that liquid transmits, and has just formed pump.The valve film on the inverted order operation left side and the right can change the direction that fluid flows, and realizes the two-way operation of pump.Among Fig. 9 up and down arrow represent to act on the direction of the power of barrier film; Right arrow is represented the pumping direction.Before operation, pump is in a state, and all valves all are in closed condition.When the b state, the feed liquor opening of valves, liquid is inhaled into the feed liquor valve from liquid inlet channel.Next be the c state, pumping diaphragm is opened, and is drawn into more liquid and enters pumping system.D, e, f state are respectively: the feed liquor valve cuts out, and liquid valve is opened, and pumping diaphragm closes to pump liquid.
Wherein, described micro-fluidic control module also comprises the analytical work platform, and the micro-fluidic chip seat is arranged on the platform, reaches fixedly fastener of chip, and chip stably is fixed on the chip carrier.
Wherein, described fastener is mechanical type fastener or vapour-pressure type fastener.
Wherein, described detecting unit comprises excitation source and excitation light path unit thereof, receiving light path unit, signal processing unit.
Using device provided by the invention to carry out the method for micro-fluidic time resolved fluoro-immunoassay, is on micro-fluidic chip, and by the control to the microfluidic valve pump, the institute that deadline resolution fluorescence immunoassay detects in steps.
Particularly, micro-fluidic time-resolved fluorescence immunoassay method may further comprise the steps:
(1) sample is added sample pool, dilution adding dilution liquid pool, cleaning fluid adds the cleaning liquid pool, and marking fluid adds the mark liquid pool, and the enhancing liquid that dissociates adds the enhancing liquid pool that dissociates;
(2) start micro-fluidic control module work: the antigen-antibody combination, effluent discharge is cleaned, effluent discharge, the mark combination, effluent discharge is cleaned, effluent discharge, the enhancing of dissociating detects.
Wherein, antigen-antibody in conjunction with detailed process is described in the step (2): sample pool-dilution liquid pool two-way pump begins two-way operation, and sample is mixed with dilution; Mixed liquor all is stored in sample pool when stopping.
Described effluent discharge detailed process is: sample pool-waste liquid pool one-way pump one-way only operation enters waste liquid pool with mixed liquor.
Described cleaning detailed process is: sample pool-cleaning liquid pool one-way pump one-way only operation sucks sample pool with cleaning fluid.
Perhaps, described cleaning detailed process is: as cleaning Buffer Pool, sample cell is got rid of waste liquid earlier, sucks cleaning fluid, closes one-way pump sample scavenging pump and one-way pump sample waste drains pump then with the dilution liquid pool; Start two-way pump, cleaning fluid is flowed between sample pool and dilution liquid pool, clean; Cleaning fluid is in sample pool when stopping.
Described mark in conjunction with detailed process is: two-way pump starts, and marking fluid is flowed between sample pool and mark pond, carries out the mark combination, stops tense marker liquid in sample cell.
The described enhancing detailed process of dissociating is: the two-way pump startup, and making the enhancing liquid that dissociates at sample pool and dissociating to strengthen between liquid pool and flowing, dissociate, dissociate enhancing liquid when stopping in the enhancing liquid pool that dissociates.
Described detection detailed process is: detecting unit moves to the detection position of micro-fluidic detection chip and detects.
The application of method in biomedical research or clinical ultramicron biochemical investigation that the present invention proposes.
Beneficial effect of the present invention is:
1. the employing microflow control technique is simplified detection system; Saved the microwell plate oscillator, microwell plate is washed the plate machine, and micro-fluidic control system is simple in structure, optimizes, and mechanical motion and structures such as no cleaning, vibration, transmission are convenient to realize robotization;
2. microflow control technique has improved reaction efficiency: the antigen-antibody combination, in sample pool, antibody sandwich is at sample pool, membrane pump work, solution is ceaselessly back and forth flowed between two ponds, wash away sample pool inwall bag quilt cover, make antigen or antibody in the solution effectively contact more abundant with antibody or the antigen of bag quilt cover;
3. improved cleaning efficiency: micro-fluidic pump valve drives liquid flow, realizes flowing and cleans;
4. detection speed improves;
5. the preparation of sample and reagent before reaction, in separately container in sample and the disposable adding chip of all ingredients, is finished sample, reagent is prepared; The operation that sample, reagent are carried out is afterwards undertaken by micro-fluidic, in the sample with a collection of detection, and all operation control, it is on all four reaching the reaction time, has avoided the deviation on the reaction time between sample;
6. sample and reagent are communicated with mixing, cleaning by the valve pump of micro-fluidic inside, the sputter pollution of having avoided using pipettor, sample injector moves may cause above chip;
7. mark combination, dissociation process and antigen-antibody cohesive process are similar, and be same because membrane pump drives flow of solution, improved reaction velocity;
8. the pollution to personnel and environment has been avoided in operating process sealing, improves the experiment biological safety.
In a word, micro-fluidic time resolved fluoro-immunoassay device of the present invention adopts microflow control technique, and detection system is simplified; Saved the microwell plate oscillator, microwell plate is washed the plate machine, and micro-fluidic control system is simple in structure, optimizes, and mechanical motion and structures such as no cleaning, vibration, transmission are convenient to realize robotization; Micro-fluidic time-resolved fluorescence immunoassay method of the present invention has improved the detection efficiency of sample greatly.
Description of drawings
Fig. 1 is the micro-fluidic time resolved fluoro-immunoassay structure drawing of device of the present invention;
Fig. 2 is micro-fluidic chip vertical view of the present invention;
Fig. 3 is micro-fluidic chip of the present invention bottom upward view;
Fig. 4 is sample pool of the present invention-dilution liquid pool two-way pump workflow diagram;
Fig. 5 is sample pool of the present invention-waste liquid pool one-way pump and sample pool-cleaning liquid pool one-way pump workflow diagram;
Fig. 6 dilutes liquid pool and cleans the Buffer Pool workflow diagram for the present invention;
Fig. 7 is sample pool of the present invention-mark liquid pool two-way pump workflow diagram;
Fig. 8 is sample pool-dissociation solution of the present invention (detection) pond two-way pump workflow diagram;
Fig. 9 is membrane pump mechanical work principle figure.
Wherein, it is as follows to install each parts sequence number:
Embodiment
Following examples are used for explanation the present invention, but are not used for limiting the scope of the invention.Without departing from the spirit and substance of the case in the present invention, modification or replacement to the inventive method, step or condition are done all belong to protection scope of the present invention.
If do not specialize the conventional means that used technological means is well known to those skilled in the art among the embodiment.
Embodiment 1: micro-fluidic time resolved fluoro-immunoassay device
As Fig. 1, Fig. 2 and shown in Figure 3, a kind of micro-fluidic time resolved fluoro-immunoassay device comprises application of sample unit 101, micro-fluidic control module 103, detecting unit 104, and it also comprises the micro-fluidic chip 102 that adopts the membrane pump type of drive.Micro-fluidic chip 102 is arranged in the micro-fluidic chip seat on the micro-fluidic control module 103 analytical work platforms.
Wherein, sample pool 201 inwalls are coated with antibody or antigen.
This micro-fluidic chip 102 adopts light tight polymeric material polystyrene (PS) preparation.
Embodiment 2: with the measurement device hepatitis b virus s antigen of embodiment 1
1, coated antibody: antibody is anti-hepatitis B virus surface antigen (anti-HBs) monoclonal antibody
1) sodium carbonate-sodium bicarbonate buffer liquid with 50mM pH9.6 will doubly dilute for the antibody 1:6000 of bag quilt, and is standby;
2) antibody with dilution adds in the sample pool by 100 μ l/ holes;
3) sample pool places 4 ℃ of environment, and bag was by 20 hours;
4) bag is moved to end, and by 300 μ l/ holes, with washing lotion washing 2 times, the washing lotion composition is that the PBS of 10mMpH7.4 contains 5% Tween-20 (volume ratio);
5) washing finishes, and sample pool is placed blot surplus solution on the absorbent filter, and the amount by 150 μ l/ holes adds confining liquid in sample pool afterwards, and sealing is 2 hours under the room temperature, and the confining liquid composition is that the PBS of 10mM pH7.4 contains the 1%BSA(bovine serum albumin);
6) sealing finishes, and confining liquid is thrown away, and sample pool is placed blot surplus solution on the absorbent filter, places 28 ℃ of constant incubator inner dryings then 20 hours.
2, labelled antibody:
1) Eu
3+The preparation of-DTPA marking fluid
(contain Eu with purified water
3+10
-6Mol/L) dilution 1-(4-isothiocyanic acid benzyl) diethylene triamine pentacetic acid (DTPA) (being called for short DTPA) places 37 ℃ of waters bath with thermostatic control to add thermal response 2 hours the solution after the dilution;
2) Eu
3+-DTPA labelled antibody
1mg antibody to 4 ℃ of dialysis of 0.1M carbonate buffer solution (pH9.3) 16 hours, is transferred to the EP pipe with antibody-solutions after the dialysis, gets Eu
3+-DTPA 0.2mg adds in the antibody-solutions, and the room temperature lucifuge stirred 14 hours;
3) purifying
Superdex 200 filler mixings are packed in 1 * 30cm post, treat that filler sinks after, carry out compression leg with purified water, flow velocity is controlled at 3ml/min, flows 2 column volumes and gets final product.After pillar presses, handle with the pillar of 0.1mmol/L NaOH, flow velocity is controlled at 3ml/min, and 2 column volumes get final product.Wash with water flatly then, use column equilibration liquid (0.1% high-purity BSA aqueous solution) to purification column balance 1 hour again.Draw the antibody that mark is good with pipettor and slowly join in the pillar, with eluent (50mMTris-HCl contains 0.9%NaCl and 0.05% Sodium azide pH7.8) sample is carried out wash-out, flow velocity is controlled at 1ml/min.
4) collect
Sample 1ml/ pipe behind the collection wash-out, select five high pipes of absorbance to merge according to the absorbance of Protein Detection instrument 280nm albumen, and with the sample that merges through 0.22 μ m membrane filtration degerming, place 4 ℃ of environment to preserve, obtain the antibody-solutions of europium mark, be called for short the europium marking fluid.
3, preparation is dissociated and is strengthened liquid, dilution, cleaning fluid
Enhancing liquid dissociates: the 6ml glacial acetic acid Potassium Hydrogen Phthalate adjust pH 3.2 of 0.1M, add 15 μ mol β-NTA(β-naphthoyltrifluoroacetones), 50 μ mol TOPO(trioctyl-phosphine oxide), 1ml Triton X-100, purified water is settled to 1L, stirs and evenly mixs.
Sample dilution: contain 1% bovine serum albumin(BSA), contain the Tris-HCl damping fluid of 0.02% disodium ethylene diamine tetraacetate;
Cleaning fluid: the 0.2MTris-HCl damping fluid that contains 5%Tween20 (polysorbas20);
4, detection method:
1) referring to Fig. 1, Fig. 2.100 μ l samples (clinical sample) are joined in micro-fluidic detection chip 102 sample pools 201, micro-fluidic detection chip 102 is placed on the micro-fluidic control module 103, place dilution, cleaning fluid, marking fluid, enhancing liquid dissociates;
2) start detection, application of sample unit 101(self-control) 200 μ l dilutions are added dilution liquid pool 202, the 2.0ml cleaning fluid adds and cleans liquid pool 205,200 μ l europium marking fluids and add mark liquid pool 203,200 μ l and dissociate and strengthen liquid and add to dissociate and strengthen liquid pool 204;
3) antigen-antibody (bag quilt) reaction bonded; Sample pool 201-dilution liquid pool 202 two-way pumps 402 beginning two-way operations (Fig. 4, Fig. 6) make sample mix with dilution, continue 50min; Mixed liquor all is stored in sample pool 201 when stopping;
4) effluent discharge, 406 one-way only operations of sample pool 201-waste liquid pool 206 one-way pumps enter waste liquid pool 206 with mixed liquor;
5) suck cleaning fluid, sample pool 201-cleans liquid pool 205 one-way pumps, 405 one-way only operations (Fig. 5), and cleaning fluid is sucked sample pool 201;
6) clean repeated execution of steps 4) effluent discharge, 5) suck cleaning fluid, carry out 4 times;
7) europium mark combination, two-way pump 403 starts, and 150 μ l europium marking fluids are flowed (Fig. 7) at sample pool 201 and 203 in mark pond, and " tape label antibody "+" antigen-antibody (bag quilt) reaction bonded body forms sandwich " continues about 30min; Stop tense marker liquid in sample cell 201, execution in step 4 then) get rid of waste liquid in waste liquid pool 206;
8 wash plate, execution in step 6) clean;
9) enhancing of dissociating, two-way pump 404 starts, and 150 μ l is dissociated strengthen liquid at sample pool 201 and dissociate to strengthen 204 of liquid pools and flow, and dissociates, and continues 5min, dissociates when stopping to strengthen liquid and strengthen in the liquid pool 204 (Fig. 8) dissociating;
Above-mentioned steps needs time 110-120min.
After adding dissociation solution, europium ion can disintegrate down from forming sandwich labelled antibody with coated antibody, produces fluorescence through exciting the back, according to the light intensity that excites the back to produce, can determine the concentration of hepatitis B surface antigen.With the hepatitis B surface antibody standard items (15ng/ml) of quadrat method test concentration known, parallel detection 20 holes, its precision CV＜7% with the comparison of classic method, can improve detection precision, dwindles the coefficient of variation.According to " Chinese biological goods rules 2000 " version regulation, hepatitis B surface antibody diagnostic kit precision CV≤15%.
Table 1: parallel detection 20 hole sample results
Embodiment 3: with the measurement device hepatitis b virus s antigen of embodiment 1
In the step 4, cleaning process is: sample pool-cleaning liquid pool one-way pump 405 one-way only operations sucks sample pool 201 with cleaning fluid.
As cleaning Buffer Pool, sample cell 201 is got rid of waste liquid earlier, sucks cleaning fluid, closes one-way pump sample scavenging pump 405 and one-way pump sample waste drains pump 406 then with dilution liquid pool 202; Start two-way pump 402, cleaning fluid is flowed at sample pool 201 and 202 of dilution liquid pools, clean; Cleaning fluid is in sample pool 201 when stopping.
Other step is identical with embodiment 2.
The above only is preferred implementation of the present invention; should be pointed out that for those skilled in the art, under the prerequisite that does not break away from the technology of the present invention principle; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (12)
1. micro-fluidic time resolved fluoro-immunoassay device, comprise application of sample unit (101), micro-fluidic control module (103), detecting unit (104), it is characterized in that, it also comprises the micro-fluidic chip (102) that adopts the membrane pump type of drive, and described micro-fluidic chip (102) is arranged in the micro-fluidic chip seat on micro-fluidic control module (103) the analytical work platform.
2. device as claimed in claim 1, it is characterized in that described micro-fluidic chip (102) comprising: sample pool (201), dilution liquid pool (202), mark liquid pool (203), the liquid pool that dissociates (204), cleaning liquid pool (205), waste liquid pool (206);
Be provided with sample valve (301) and sample through hole (211) in the wherein said sample pool (201); Be provided with dilution valve (302) and dilution through hole (212) in the dilution liquid pool (202); Be provided with marking fluid valve (303) and marking fluid through hole (213) in the described mark liquid pool (203), be provided with dissociation solution valve (304) and the fluid through-hole that dissociates (214) in the described liquid pool that dissociates (204), described cleaning liquid pool (205) is provided with cleaning fluid valve (305) and cleaning fluid through hole (215); Described waste liquid pool (206) is provided with waist valve (306) and waste liquid through hole (216);
Main valve (307) is arranged at the centre of each valve, and each valve links to each other with main valve (307) respectively by passage.
3. device as claimed in claim 2 is characterized in that, described sample valve (301), main valve (307) and dilution liquid pool valve (302) and passage constitute sample pool (201)-dilution liquid pool (202) two-way pump sample dilution pumps (402); Described sample valve (301), main valve (307) and waist valve (306) and passage constitute sample pool (201)-waste liquid pool (206) one-way pump sample waste drains pump (406); Described sample valve (301), main valve (307) and cleaning fluid valve (305) and passage constitute sample pool (201)-cleaning liquid pool (205) one-way pump sample scavenging pump (405); Described sample valve (301), main valve (307) and marking fluid valve (303) and passage constitute sample pool (201)-mark liquid pool (203) two-way pump sample labeling pump (403); Described sample valve (301), main valve (307) and dissociation solution valve (304) and passage constitute sample pool (the 201)-liquid pool that dissociates (204) two-way pump sample solution from pump (404).
4. as any described device of claim 1-3, it is characterized in that described sample pool (201) inwall is coated with antibody or antigen.
5. as any described device of claim 1-3, it is characterized in that described micro-fluidic chip (102) is made for light tight polymeric material.
6. application rights requires the arbitrary described device of 1-5 to carry out the method for micro-fluidic time resolved fluoro-immunoassay, it is characterized in that, may further comprise the steps:
1) sample is added sample pool (201), dilution adding dilution liquid pool (202), liquid pool (205) is cleaned in the cleaning fluid adding, and marking fluid adds mark liquid pool (203), and the enhancing liquid that dissociates adds the enhancing liquid pool (204) that dissociates;
2) start micro-fluidic control module (103) work: the antigen-antibody combination, effluent discharge is cleaned, effluent discharge, the mark combination, effluent discharge is cleaned, effluent discharge, the enhancing of dissociating detects.
7. method as claimed in claim 6 is characterized in that step 2) described in antigen-antibody in conjunction with detailed process be: (202) two-way pump sample dilution pumps (402) two-way operation that sample pool (201)-dilution liquid pool is formed makes sample mix with dilution;
Step 2) detailed process of effluent discharge described in is: one-way pump sample waste drains pump (406) one-way only operation that sample pool (201)-waste liquid pool (206) is formed enters waste liquid pool (206) with mixed liquor.
8. method as claimed in claim 6 is characterized in that step 2) described in clean detailed process and be: one-way pump sample scavenging pump (405) one-way only operation of sample pool (201)-cleanings liquid pool (205) composition sucks sample pool (201) with cleaning fluid.
9. method as claimed in claim 6, it is characterized in that, step 2) cleaning detailed process described in is: to dilute liquid pool (202) as cleaning Buffer Pool, sample cell (201) is got rid of waste liquid earlier, suck cleaning fluid, close one-way pump sample scavenging pump (405) and one-way pump sample waste drains pump (406) then; Start two-way pump (402), cleaning fluid is flowed between sample pool (201) and dilution liquid pool (202), clean; Cleaning fluid is in sample pool (201) when stopping.
10. method as claimed in claim 6, it is characterized in that step 2) described in mark in conjunction with detailed process be: two-way pump (403) starts, and makes marking fluid mobile between sample pool (201) and mark pond (203), carry out the mark combination, stop tense marker liquid in sample cell (201).
11. method as claimed in claim 7, it is characterized in that, step 2) the enhancing detailed process of dissociating described in is: two-way pump (404) starts, making dissociates strengthens liquid at sample pool (201) and dissociates mobile between enhancing liquid pool (204), dissociate, dissociating when stopping to strengthen liquid in the enhancing liquid pool (204) that dissociates.
12. the application of any described device of claim 1-5 in biomedical research or clinical ultramicron biochemical investigation.
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| CN104897635A (en) * | 2015-06-19 | 2015-09-09 | 上海凯创生物技术有限公司 | Immunofluorescence analyzer |
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| CN106338601A (en) * | 2016-08-26 | 2017-01-18 | 周辉 | United and quantitative anti-sperm antibody and Mullerian duct hormone detection card and manufacturing method and usage method thereof |
| CN107321397A (en) * | 2017-05-31 | 2017-11-07 | 深圳市海拓华擎生物科技有限公司 | A kind of micro-fluidic chip and its application |
| CN108080042A (en) * | 2017-11-13 | 2018-05-29 | 成都微康生物科技有限公司 | Micro-fluidic chip of binding time resolved fluorometric technology and its preparation method and application |
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| CN117471105A (en) * | 2023-12-27 | 2024-01-30 | 北京芯迈微生物技术有限公司 | Microfluidic chip for bidirectional double-drive sequential platelet HLA antibody detection and application |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100303687A1 (en) * | 2009-06-02 | 2010-12-02 | Integenx Inc. | Fluidic devices with diaphragm valves |
| CN102166537A (en) * | 2011-01-30 | 2011-08-31 | 南京大学 | Hydrophilic, multifunctional and integrated miniflow control chip easy to optical detection, manufacture method thereof and use thereof |
| CN102851199A (en) * | 2012-10-09 | 2013-01-02 | 重庆大学 | Micro-fluidic cell sorting and acquiring device |
-
2013
- 2013-03-14 CN CN201310081065.9A patent/CN103207169B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100303687A1 (en) * | 2009-06-02 | 2010-12-02 | Integenx Inc. | Fluidic devices with diaphragm valves |
| CN102166537A (en) * | 2011-01-30 | 2011-08-31 | 南京大学 | Hydrophilic, multifunctional and integrated miniflow control chip easy to optical detection, manufacture method thereof and use thereof |
| CN102851199A (en) * | 2012-10-09 | 2013-01-02 | 重庆大学 | Micro-fluidic cell sorting and acquiring device |
Non-Patent Citations (1)
| Title |
|---|
| 马冬红等: "血清促甲状腺素免疫检测技术", 《上海交通大学学报(医学版)》 * |
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| CN104897635B (en) * | 2015-06-19 | 2017-06-16 | 上海凯创生物技术有限公司 | Immunofluorescence analysis instrument |
| CN105381827A (en) * | 2015-12-17 | 2016-03-09 | 北京博晖创新光电技术股份有限公司 | Microfluidic chip magnetic bead even mixing apparatus and method |
| CN106338601A (en) * | 2016-08-26 | 2017-01-18 | 周辉 | United and quantitative anti-sperm antibody and Mullerian duct hormone detection card and manufacturing method and usage method thereof |
| CN106338601B (en) * | 2016-08-26 | 2019-04-05 | 广东普洛宇飞生物科技有限公司 | A kind of AsAb and anti-Miao Leguan hormons quantitative test card and its production, application method |
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| CN110672829A (en) * | 2019-09-10 | 2020-01-10 | 中国科学院上海技术物理研究所 | Microfluidic laser immunity time domain spectrum detection method |
| CN110672828A (en) * | 2019-09-10 | 2020-01-10 | 中国科学院上海技术物理研究所 | Rare animal blood detector of micro-fluidic laser immunity time domain spectrum |
| CN117471105A (en) * | 2023-12-27 | 2024-01-30 | 北京芯迈微生物技术有限公司 | Microfluidic chip for bidirectional double-drive sequential platelet HLA antibody detection and application |
| CN117471105B (en) * | 2023-12-27 | 2024-03-26 | 北京芯迈微生物技术有限公司 | Microfluidic chip for bidirectional double-drive sequential platelet HLA antibody detection and application |
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