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Small molecule triple immunochromatography detection method, test strip and kit

An immunochromatographic detection and small molecule technology, applied in analytical materials, biological testing, measurement devices, etc., can solve the problems of many influencing conditions, limited detection sensitivity, difficult to control accurately, etc., to improve detection sensitivity and wide range of detection applications. Effect

Pending Publication Date: 2021-01-05
天津浩泰科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this type of test strip technology has certain shortcomings: 1. In the double detection, the detection range is difficult to connect, which is not conducive to data analysis; the triple detection technology has not been reported; 2. The antigen immobilization of the test substance is not conducive to the full exposure of the binding site. It is not conducive to the full progress of the competitive binding reaction, which limits the detection sensitivity; 3. The competitive binding reaction that occurs in the detection area of ​​the chromatography test paper affects many conditions and is difficult to accurately control, which is not conducive to the detection sensitivity.

Method used

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  • Small molecule triple immunochromatography detection method, test strip and kit
  • Small molecule triple immunochromatography detection method, test strip and kit
  • Small molecule triple immunochromatography detection method, test strip and kit

Examples

Experimental program
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Effect test

Embodiment 1

[0055] This embodiment takes the detection of diazepam as an example to illustrate the preparation of the detection test strip of the present invention. The modified protein was selected from BSA, and the signal substance of the labeled antibody was selected from carboxylated fluorescent quantum dot microspheres (QB, purchased from Beijing Najing Biotechnology Co., Ltd., product model: QBB12117, H07187M).

[0056] The labeled antigen signal substance is selected from the Raman signal surface enhancement material Au@Ag-R, which is prepared by the following method: reducing chloroauric acid to gold seeds by sodium citrate, further reducing silver nitrate by sodium citrate reduction method and coating on Gold seeds to realize the preparation of Au@Ag. After the preparation of the material is completed, a Raman reporter molecule dissolved in 5% methanol aqueous solution is added at a concentration of 10 mg / mL; the Raman reporter molecule is DTNB: 5,5'-dithiobis(2-nitrobenzoic acid...

Embodiment 2

[0064] The present embodiment takes the detection of diazepam as an example to illustrate the detection method of the present invention, test with the test strip that embodiment 1 makes:

[0065] Prepare standard curve

[0066] Diazepam (DAP) standard was prepared as DAP solutions with different concentrations in phosphate buffer: 1.28ng / mL, 0.64ng / mL, 0.32ng / mL, 0.16ng / mL, 0.08ng / mL, 0.04ng / mL, 0.02ng / mL, 0.01ng / mL. The composition of the phosphate buffer is as follows: based on 0.05mol / L phosphate buffer, the following substances are added in the following weight percentages: 2% sucrose, 5% fructose, 1% PEG, 3% Tween-20.

[0067] Au@Ag-R-DAP-BSA 0.1μg / 100μL solution, QB-DAP-mAb were mixed with different concentrations of DAP standards at a dilution ratio of 1:3000, QB-DAP-mAb and Au@Ag-R-DAP -The final concentration range of BSA is 0.2-0.8mg / mL and 0.01-0.04mg / mL respectively. At 37℃, DAP was incubated with labeled antibody QB-DAP-mAb and Au@Ag-R-DAP-BSA for 10min, A mix...

Embodiment 3

[0073] This embodiment takes DAP in aquatic products as an example to illustrate that the sample to be tested is detected by the method of the present invention, and the steps are as follows:

[0074] Sample to be tested: DAP was added to fish meat;

[0075] The test strip prepared in Example 1 was equilibrated to room temperature.

[0076] The fish meat spiked with DAP was diluted with a mixed solution of phosphate buffer saline and methanol (10%) in equal volume, and the resulting diluted solution was subjected to the immunochromatographic process.

[0077] The diluted fish meat standard sample was diluted and incubated with labeled antibody and labeled antigen for 11 minutes at 37°C to form immune complexes QB-DAP-mAb-DAP and QB-DAP-mAb-Au@Ag-R-DAP -BSA mixture; after incubation, the solution containing the mixture is added to the glass fiber pad, and through the suction of the water-absorbing pad, it passes through the detection area and the quality control area of ​​the re...

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Abstract

The invention belongs to the field of small molecule detection, and relates to a small molecule triple immunochromatography detection method, a test strip and a kit. The detection method comprises thesteps of (1) incubating an object to be detected, a labeled antibody and a labeled antigen to obtain a mixture, wherein the labeled antibody is an antibody for resisting target small molecules and islabeled with a first signal substance, the labeled antigen is a derivative of a target small molecule with a modified protein and is labeled with a second signal substance; (2) enabling the mixture to sequentially pass through a detection area and a quality control area, wherein an antibody for resisting the modified protein in the labeled antigen is fixedly arranged in the detection area, and asecondary antibody corresponding to the labeled antibody is fixedly arranged in the quality control area; and (3) obtaining the content of the target small molecules in the to-be-detected object according to the triple signal intensity of the detection area and the quality control area. The detection method is sensitive, fusion of triple detection can be achieved, a wider detection range is provided for small molecule detection, and the detection method has wide market prospects.

Description

technical field [0001] The invention belongs to the field of small molecule detection, and in particular relates to a small molecule triple immunochromatography detection method, a small molecule triple immunochromatography detection test strip and a small molecule triple immunochromatography detection kit. Background technique [0002] There are many existing detection methods for small molecules. Among them, the detection principle of the immunochromatographic test paper based on the principle of chromatographic separation is to pre-fix the specific antigen of the test object to the test area of ​​the test paper. Under the action of capillary force, the test paper in the sample The small molecules and labeled antibodies flow to the measurement area, and the immobilized antigen competes with the analytes in the solution to bind to the labeled antibodies in the solution. The signal intensity in the determination area is inversely proportional to the content of small molecule...

Claims

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Application Information

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IPC IPC(8): G01N33/558G01N33/58G01N33/543G01N33/53
CPCG01N33/558G01N33/543G01N33/588G01N33/585G01N33/5308G01N33/948
Inventor 张迎春苑帅刘雪飞
Owner 天津浩泰科技有限公司
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