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Method for improving sensitivity of immunochromatographic marker and application of immunochromatographic marker in interleukin 6 detection

A technology of immunochromatography and markers, which is applied in the field of immunochromatography, can solve the problems of low labeling sensitivity, achieve the effects of improving reagent sensitivity, increasing the intensity of fluorescent substances, and increasing the intensity of fluorescent signals

Pending Publication Date: 2020-06-19
SICHUAN XINCHENG BIOLOGICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Low labeling sensitivity of fluorescence chromatography is a common pain point

Method used

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  • Method for improving sensitivity of immunochromatographic marker and application of immunochromatographic marker in interleukin 6 detection
  • Method for improving sensitivity of immunochromatographic marker and application of immunochromatographic marker in interleukin 6 detection
  • Method for improving sensitivity of immunochromatographic marker and application of immunochromatographic marker in interleukin 6 detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Application of a method for improving the sensitivity of immunochromatographic markers in the detection of interleukin-6, the preparation method of the markers is as follows:

[0035] 1. Take out 500ul fluorescent microspheres (1% concentration W / V, green fluorescence-excitation 475nm-emission 525nM) into a centrifuge tube.

[0036] 2. Centrifuge (12000rpm-20000rpm according to different particle sizes) for 10min to settle the microspheres and remove the supernatant.

[0037] 3. Add 500ul coupling buffer (50mM MES pH 6.0) and mix well.

[0038] 4. Add 20ul of EDC solution (200mM), 20ul of sulfo-NHS solution (200mM), mix well, and incubate on a rotary mixer for 30min.

[0039] 5. Centrifuge (12000rpm-20000rpm according to different particle sizes) for 10min to settle the microspheres and remove the supernatant.

[0040] 6. Add 500ul coupling buffer (50mM MES pH6.0), mix well, add 0.1mg anti-IL-6 monoclonal antibody, mix well, and incubate for 1h at room temperature wit...

Embodiment 2

[0048] Application of a method for improving the sensitivity of immunochromatographic markers in the detection of interleukin-6, the preparation method of the markers is as follows:

[0049] 1. Take out 500ul fluorescent microspheres (1% concentration W / V, green fluorescence-excitation 475nm-emission 525nM) into a centrifuge tube.

[0050] 2. Centrifuge (12000rpm-20000rpm according to different particle sizes) for 10min to settle the microspheres and remove the supernatant.

[0051] 3. Add 500ul coupling buffer (50mM MES pH 6.0) and mix well.

[0052] 4. Add 20ul of EDC solution (200mM), 20ul of sulfo-NHS solution (200mM), mix well, and incubate on a rotary mixer for 30min.

[0053] 5. Centrifuge (12000rpm-20000rpm according to different particle sizes) for 10min to settle the microspheres and remove the supernatant.

[0054] 6. Add 500ul coupling buffer (50mM MES pH6.0), mix well, add 0.1mg anti-IL-6 monoclonal antibody, mix well, and incubate for 1h at room temperature with ...

Embodiment 3

[0062] Preparation of interleukin 6 (IL-6) fluorescence chromatography test strip:

[0063] The fluorescent chromatography detection test strip comprises a base plate and a sample pad, a marker pad, a chromatographic membrane and a water-absorbing pad sequentially connected on the base plate, the marker pad is a glass fiber pad, and the chromatographic membrane is a nitrocellulose membrane.

[0064] The base plate is made of PVC board, and the sticking parts between the sample pad, marker pad, chromatographic membrane and water-absorbing pad overlap each other by 2mm. After assembling and cutting, they become 4mm wide test strips, which are put into the card case and packaged in an aluminum foil bag.

[0065] Specific steps are as follows:

[0066] 1. Preparation of glass fiber mat: Cut the glass fiber into 3*3cm specifications, use a gold sprayer to spray the treatment solution (blocking agent 10%, Tween 20 0.5%, anti-erythrocyte antibody 1%, 50mM PBS ph 7.2 ) was sprayed on...

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Abstract

The invention discloses a method for improving the sensitivity of an immunochromatographic marker, which comprises the following steps of coupling an antibody with fluorescent microspheres to obtain an antibody-fluorescent microsphere conjugate, and coupling the antibody-fluorescent microsphere conjugate with the fluorescent microspheres again to obtain an enhanced antibody-fluorescent microsphereconjugate as the immunochromatographic marker. According to the method, the labeled antibody-fluorescent microsphere conjugate is coupled again, so that the same antibody can carry more fluorescent microspheres, the strength of fluorescent substances on a detection line is greatly improved, and the sensitivity of a reagent is improved. The invention also discloses an application of the method ininterleukin 6 detection.

Description

technical field [0001] The invention relates to the technical field of immunochromatography, in particular to a method for improving the sensitivity of an immunochromatography marker and its application in the detection of interleukin-6. Background technique [0002] Immunochromatography, also known as lateral flow immunoassay, originated in the 1980s and is an immunoassay technology based on the specific reaction of antigen and antibody on the chromatographic membrane. The advantages of low cost and high sensitivity are widely used in pathogen detection, food safety measurement, environmental monitoring, etc. Among them, colloidal gold immunochromatography technology is the most mature, but it is mostly limited to qualitative or semi-quantitative detection. With the vigorous development of material science and bioengineering technology, new fluorescence chromatography technology has gradually gained a firm foothold in the field of quantitative detection. In the current flu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/533G01N33/68G01N33/577
CPCG01N33/533G01N33/6869G01N33/577G01N2333/5412
Inventor 林源蒋明君李学锐李晟
Owner SICHUAN XINCHENG BIOLOGICAL CO LTD
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