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Magnetic particle chemiluminescence micro-fluidic chip used for whole-blood sample detection

A microfluidic chip and chemiluminescence technology, which is applied in chemiluminescence/bioluminescence, analysis through chemical reaction of materials, biological testing, etc., can solve the problems of low sensitivity, interference, poor repeatability, etc., and achieve structural Simple design, high sensitivity effect

Active Publication Date: 2016-01-13
SHENZHEN HUAMAIXINGWEI MEDICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] The technical problem to be solved by the present invention is to provide a microfluidic magnetic sensor for the problems of low sensitivity, poor repeatability, obvious interference, and the existing chemiluminescent microfluidic chip supporting equipment and long detection time of the existing rapid diagnostic method. Microparticle chemiluminescent immunoassay chip, through integrated chip (all components except the test sample are integrated into the chip) and supporting small portable equipment, so as to realize the rapid, accurate and highly sensitive quantitative detection of analytes in field samples

Method used

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  • Magnetic particle chemiluminescence micro-fluidic chip used for whole-blood sample detection
  • Magnetic particle chemiluminescence micro-fluidic chip used for whole-blood sample detection
  • Magnetic particle chemiluminescence micro-fluidic chip used for whole-blood sample detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Example 1: Double-antibody sandwich determination of hypersensitive troponin T (cTnT)

[0072] (1) Antibody labeling

[0073] Dissolve 5 μg of HRP in 1 mL of distilled water, then add 0.2 mL of 0.1 M freshly prepared NaIO 4 The solution was reacted at room temperature in the dark for 20 minutes, and the purified solution was dialyzed against 1 mM pH 4.4 sodium acetate buffer. Then adjust the pH to 9.0 with 0.2 M pH9.5 carbonate buffer, add 10 μg of anti-cTnT monoclonal antibody, and react at room temperature for 2 h in the dark. Add 0.1mL freshly prepared 4mg / mLNaBH 4 solution, mixed well, and reacted at 4°C for 2h. The above solution was put into a dialysis bag, dialyzed against 0.15M pH7.4PBS, overnight at 4°C, and HRP-labeled cTnT antibody was obtained.

[0074] Add 1mg magnetic particles (2μm in size), 10μg EDC and 15μg NHS solution and 15μg anti-cTnT monoclonal antibody (different from HRP-labeled antibody) solution to 1ml 10mM pH7.4 phosphate buffer, mix well ...

Embodiment 2

[0085] Embodiment 2: Determination of tacrolimus blood drug concentration by competitive method

[0086] (1) Antibody / antigen labeling

[0087] Weigh 0.1 mg of ALP and dissolve it in 0.1 ml of 12.5% ​​glutaraldehyde pH 6.80.1 MPBS solution, overnight at room temperature. The reacted enzyme solution was passed through a SephadexG-25 chromatographic column, eluted with physiological saline, and the brown liquid was collected. With stirring, 22.5 μg of anti-tacrolimus antibody was added dropwise into the enzyme solution, and reacted for 3 hours. Add 0.2M lysine to block, mix well, and place at room temperature for 2h. Add an equal volume of saturated ammonium sulfate dropwise with stirring, and place at 4°C for 1h. Centrifuge at 3000rpm / min for 30min, discard the supernatant. The precipitate was washed twice with half-saturated ammonium sulfate, and finally the precipitate was dissolved in 0.15 M pH 7.4 PBS. Put the above solution into a dialysis bag, dialyze against 0.15MpH...

Embodiment 3

[0101] Example 3: Magnetic Particle Size Screening

[0102] For other experimental conditions, refer to Example 1. The size of the magnetic particles and the magnetic induction of the magnet are carried out according to the following scheme.

[0103] The particle size is 0.1 μm, 0.5 μm, 1.1 μm, 1.5 μm, 2.5 μm, 3 μm, 10 μm. The magnetic induction of the magnet is 500 Gauss, 1000 Gauss, 4000 Gauss, 8000 Gauss, 12000 Gauss, 30000 Gauss. Magnetic particles of seven sizes are respectively driven by the six kinds of magnets.

[0104] The experimental results show that when 0.1μm magnetic particles are combined with a 500 Gauss magnet, the minimum detection limit is 50pg / ml, the quantitative detection range is 50-5000pg / ml, and the linear correlation coefficient R 2 >0.90; both the intra-assay and inter-assay repeatability are less than 20%. That is: the chemiluminescent signal is weak, the sensitivity is not high, and the repeatability is poor.

[0105] When 10μm magnetic partic...

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Abstract

The invention discloses a magnetic particle chemiluminescence micro-fluidic chip used for whole-blood sample detection. The chip is composed of top adhesive tape (12), a chip substrate (1) and bottom adhesive tape (15), wherein a filtering area (2), a coating area (3), a reacting area (5), a cleaning area (6), a detecting area (7) and liquid release channels (8) on the chip substrate are sequentially connected, a marking ligand storage pool (4) on the chip substrate is connected with the reacting area (5), the detecting area (7) is connected with a cleaning liquid storage pool (9) and a light-emitting substrate liquid storage pool (10) through the liquid release channels (8), and the top adhesive tape comprises a sample adding port (13). Magnetic particle marking ligands are coated with the coating area (3). The marking ligand storage pool (4) is used for storing enzyme or light-emitting agent marking ligands.

Description

technical field [0001] The invention relates to a chip for highly sensitive quantitative detection of analytes using magnetic particle chemiluminescence technology and microfluidic chip technology, and discloses a magnetic particle chemiluminescence microfluidic chip for whole blood sample detection, which can realize The accurate and highly sensitive quantitative detection of pathogens, major diseases (such as tumors, cardiovascular diseases), illegal drugs, drug detection, food safety and other analytes belongs to the technical field of microfluidic chip chemiluminescence immunoassay. Background technique [0002] At present, there are two main development trends for in vitro diagnostics (IVD): one is automation and integration, that is, the use of fully automated and highly sensitive large-scale instruments and equipment in the central laboratory of a large hospital to achieve high-precision disease analysis and diagnosis ; Another kind of miniaturization and bedside, tha...

Claims

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Application Information

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IPC IPC(8): G01N21/76G01N33/53
Inventor 王东李泉
Owner SHENZHEN HUAMAIXINGWEI MEDICAL TECH CO LTD
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