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Saliva-anti-mitochondrial-antibody-M2-type enzyme-linked immunity detection kit

An anti-mitochondrial antibody and enzyme-linked immunoassay technology, which is applied in the fields of biotechnology and medical testing, can solve problems such as the absence of kits and methods, high cost of blood sample processing, and discomfort of subjects, so as to provide data Support, improve patient prognosis, and aid in diagnosis

Inactive Publication Date: 2016-11-23
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the existing blood testing technology is very mature, it has certain limitations: 1. First, the acquisition of blood is an invasive method, and the acquisition of samples requires professional skills; 2. During the blood acquisition process, the affected Test subjects are prone to discomfort and a certain risk of infection; 3. The cost of obtaining and processing blood samples is relatively high
Currently, the detection of AMA-M2 is limited to the blood route, and the kits and methods for AMA-M2 detection using saliva have not yet appeared

Method used

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  • Saliva-anti-mitochondrial-antibody-M2-type enzyme-linked immunity detection kit
  • Saliva-anti-mitochondrial-antibody-M2-type enzyme-linked immunity detection kit
  • Saliva-anti-mitochondrial-antibody-M2-type enzyme-linked immunity detection kit

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Test kit sensitivity:

[0040] A: Dilute the AMA-M2 standard with the sample diluent (0.1% BSA-PBS solution) doubly diluted (1:2 volume ratio) into 8 gradients, so that the AMA-M2 content is:

[0041] Standards of 1RU / ml, 0.50RU / ml, 0.25RU / ml, 0.125RU / ml, 0.0625RU / ml, 0.03175RU / ml, 0.015625RU / ml, ORU / ml; the 8th (ORU / ml) Set as a blank control well, directly as the sample diluent.

[0042] B: Antigen-antibody reaction, add 100 ul diluted standard substance (two columns of standard substance, each set 1 complex empty) in the polystyrene ELISA microwell plate coated with the M2 antigen, after incubating at room temperature for 30 minutes, Discard the liquid in the wells, add 300ul to each well to prepare the washing solution, pour out the liquid in the microwell plate after 60 seconds, and pat on absorbent paper to remove the residual liquid, and repeat the washing 3 times.

[0043] C: Add 100ul of the enzyme-labeled secondary antibody to each well, incubate at room tem...

Embodiment 2

[0053] One, utilize the method for detecting the AMA-M2 level in saliva of kit of the present invention, comprise the following steps:

[0054] Saliva samples were obtained from people participating in the medical examination.

[0055] This step specifically includes:

[0056] A: Before the saliva collection, the subjects were asked to rinse their mouths with plenty of water, and the saliva collection began 5 minutes after the gargle was completed. B: Ask the subject to lie flat and open the mouth, and the gag is fixed, so that the saliva flows out naturally and gathers at the bottom of the mouth. After about 60 seconds, suck the saliva in the mouth into a 15ml centrifuge tube with an aspirator. Repeat several times to get 1-3ml of saliva.

[0057] C: Centrifuge the saliva at 4°C, 500g, 10min. After centrifugation, absorb the supernatant and put it in a 1.5ml centrifuge tube for later use.

[0058] 2. Detection of AMA-M2 levels in saliva by enzyme-linked immunosorbent assa...

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Abstract

The invention relates to the field of biotechnology and medical detection, and aims at providing a saliva-anti-mitochondrial-antibody-M2-type enzyme-linked immunity detection kit. The kit is prepared from an AMA-M2 standard substance, a polystyrene ELISA micro-pore plate coated with an M2 antigen, a horseradish peroxidase-conjugated antibody, a sample diluent and auxiliary reagent, wherein the enzyme-labeled second antibody is anti-human IgG labeled by horseradish peroxidase; the sample diluent is a BSA-PBS solution with the mass concentration of 0.1%; the auxiliary reagent comprises substrate color developing liquid, reaction stopping liquid and washing damping liquid. The kit is easier and more convenient to use, the expression level of AMA-M2 in saliva of a physical examination person can be directly detected, a wound-free detection mean is achieved, and the defects exist in blood path detection are overcome. The saliva-anti-mitochondrial-antibody-M2-type enzyme-linked immunity detection kit can be more safely, conveniently and rapidly applied to screening of PBC and assisting diagnosis of PBC, and data supporting is provided for treatment of later disease and improving prognosises of a patient.

Description

technical field [0001] The invention relates to the fields of biotechnology and medical detection, in particular to an enzyme-linked immunoassay kit for salivary anti-mitochondrial antibody M2 type, aiming to establish a technology for rapid and non-invasive detection of AMA-M2 levels in human body. Background technique [0002] Enzyme-linked immunosorbent assay (ELISA) is one of the more mature technologies in the field of analytical chemistry, and it is an immunoassay technology developed on the basis of immunoenzyme technology. This technology uses the specific reaction of antigen and antibody to connect the analyte to the enzyme, and then produces a color reaction through the enzyme and the substrate for quantitative determination. The object of measurement may be an antibody or an antigen. When measuring, the antigen (antibody) is first bound to the solid phase carrier, but still retains its immunological activity, and then a conjugate (marker) formed by combining the ...

Claims

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Application Information

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IPC IPC(8): G01N33/543
CPCG01N33/543
Inventor 刁宏燕卢翀陈佳宁侯显良王琳
Owner ZHEJIANG UNIV
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