Quantum dot fluorescence chromatography immunization analysis reagent for detecting human blood ZnT8A

An immunoassay and fluorescence chromatography technology, applied in the fields of immunoassay and medical testing, can solve the problems of inability to judge the degree of pancreatic β-cell destruction, the severity of autoimmune diseases, the inability to accurately quantitatively measure autoantibodies, and the cumbersome operation methods, etc. Low, easy to operate, low cost effect

Inactive Publication Date: 2018-08-03
天津市协和医药科技集团有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The advantage of this method is that it can detect 6 kinds of antibodies at the same time, but the biggest disadvantage of western blotting is its low sensitivity and cumbersome operation method, and it can only detect autoantibodies qualitatively, and cannot accurately quantify autoantibodies in serum. Therefore, it is impossible to judge the degree of destruction of pancreatic beta cells and the severity of autoimmune diseases

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  • Quantum dot fluorescence chromatography immunization analysis reagent for detecting human blood ZnT8A
  • Quantum dot fluorescence chromatography immunization analysis reagent for detecting human blood ZnT8A

Examples

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Embodiment 1

[0041] A method for preparing a quantum dot fluorescence chromatography immunoassay reagent for detecting ZnT8A in human blood, comprising the steps of:

[0042] Preparation of sample pad: use 0.01M Tris-HCl with pH=7.2 as solvent, make anti-erythrocyte antibody into 0.15mg / ml solution, and press 35ul / cm 2 Spray the erythrocyte antibody solution onto a 1.2cm×30cm polyester fiber membrane, and dry it for later use;

[0043] Preparation of quantum dot marker-conjugated pads:

[0044] 1) Take 50ul, 10mg / mL quantum dots (the diameter of the quantum dots is 3nm), add 100ul 20mM PBS with pH=7.2, then add 15μL 25% glutaraldehyde, mix well, stir and activate at room temperature for 5 hours;

[0045] 2) Dialyze the liquid obtained in step 1) with 10-50 mM PBS at pH=7.2 for 22 hours, and change the liquid twice;

[0046] 3) Mix the mouse anti-human IgG antibody with 20mM PBS pH=7.2 to 4mg / mL, take 20μL and add it to the liquid obtained in step 2), mix well, stir and react at 5°C for 1...

Embodiment 2

[0058] A method for preparing a quantum dot fluorescence chromatography immunoassay reagent for detecting ZnT8A in human blood, comprising the steps of:

[0059] Preparation of sample pad: use 0.01M Tris-HCl with pH=7.2 as solvent, make anti-erythrocyte antibody into 0.2mg / ml solution, and press 20ul / cm 2 Spray the erythrocyte antibody solution onto the polyester fiber membrane, and let it dry for later use;

[0060] Preparation of quantum dot marker-conjugated pads:

[0061] 1) Take 50ul, 5mg / mL quantum dots (the diameter of the quantum dots is 2nm), add 100ul 20mM PBS with pH=7.2, then add 10μL 25% glutaraldehyde, mix well, stir and activate at room temperature for 6 hours;

[0062] 2) The liquid obtained in step 1) was dialyzed with 50 mM PBS with pH=7.2 for 20 hours, and the liquid was changed 3 times;

[0063] 3) Mix the mouse anti-human IgG antibody with 20mM PBS pH=7.2 to 3mg / mL, take 20μL and add it to the liquid obtained in step 2), mix well, stir and react at 2°C f...

Embodiment 3

[0074] A method for preparing a quantum dot fluorescence chromatography immunoassay reagent for detecting ZnT8A in human blood, comprising the steps of:

[0075] Preparation of sample pad: use 0.01M Tris-HCl with pH=7.2 as solvent, make anti-erythrocyte antibody into 0.1mg / ml solution, and press 50ul / cm 2 Spray the erythrocyte antibody solution onto the polyester fiber membrane, and let it dry for later use;

[0076] Preparation of quantum dot marker-conjugated pads:

[0077] 1) Take 50ul, 20mg / mL quantum dots (the diameter of the quantum dots is 4nm), add 100ul 20mM PBS with pH=7.2, then add 20μL 25% glutaraldehyde, mix well, stir and activate at room temperature for 3 hours;

[0078] 2) The liquid obtained in step 1) was dialyzed with 10 mM PBS with pH=7.2 for 24 hours, and the liquid was changed twice;

[0079] 3) Mix the mouse anti-human IgG antibody with 20mM PBS pH=7.2 to 5mg / mL, take 20μL and add it to the liquid obtained in step 2), mix well, stir and react at 8°C fo...

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Abstract

The invention discloses a quantum dot fluorescence chromatography immunization analysis reagent for detecting human blood ZnT8A. The reagent comprises a base plate, a sample pad, a quantum dot-labeledcombination pad, a chromatography reaction membrane and a water absorption pad are arranged at the upper surface of the base plate, a quantum dot-labeled mouse-anti-human IgG antibody is absorbed atthe quantum dot-labeled combination pad, a detection line fixed with ZnT8 protein and a quality control line fixed with staphylococcal protein A are provided at the chromatography reaction membrane, to-be-detected human whole blood or a blood plasma sample are dropped to the sample pad, and are subjected to a chromatography reaction for 3-4 min, a fluorescence immunoassay analyzer is used for detecting the fluorescence intensity of the detection line and the quality control line, and the zinc transportprotein 8 self-antibody content in a sample can be calculated by built-in software. The reagent has the advantages of simple operation, fast reaction, high sensitivity, and strong specificity.

Description

technical field [0001] The invention relates to the technical fields of immune analysis and medical testing, in particular, the invention relates to a quantum dot fluorescence chromatography immunoassay reagent for detecting human blood zinc transporter 8 autoantibodies and a preparation method thereof. Background technique [0002] Diabetes mellitus is a metabolic syndrome characterized by hyperglycemia, which is mainly due to absolute deficiency of insulin, resistance or tolerance of peripheral tissue cells to the biological effects of insulin, or both. Diabetes is divided into two types, type 1 and type 2, depending on the underlying cause. Type 1 diabetes is a metabolic disorder autoimmune disease characterized by the selective destruction of pancreatic β cells. It is mainly due to genetic defects that it cannot produce enough insulin independently and cannot allow cells to store excess blood sugar. Blood sugar phenomenon and cause many complications. Type 2 diabetes i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/558G01N33/68G01N33/58G01N33/533
CPCG01N33/533G01N33/558G01N33/582G01N33/588G01N33/68
Inventor 王立凯阎尔坤孙少飞郑剑锋
Owner 天津市协和医药科技集团有限公司
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