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Reagent kit for detecting autoimmunity disease related antinuclear antibodies spectrum and preparation method thereof

An autoimmune disease and anti-nuclear antibody spectrum technology, applied in the field of biomedicine, can solve problems such as misjudgment of results, destruction of antigenic determinant conformation, non-specific binding of non-specific antibodies, etc., to achieve simple operation, improved sensitivity, The effect of less sample consumption

Active Publication Date: 2008-12-24
BEIJING MOKOBIO LIFE SCI CO LTD
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  • Claims
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Problems solved by technology

However, in the process of SDS-PAGE, the conformation of many antigenic determinants may be destroyed, and they cannot be combined with specific antibodies or non-specifically, resulting in misjudgment of results

Method used

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  • Reagent kit for detecting autoimmunity disease related antinuclear antibodies spectrum and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1 Preparation of kit of the present invention

[0035] 1. Preparation of natural antigen

[0036](1) Natural antigen extraction: nRNP / Sm, Sm, SS-A, SS-B, Scl-70, Jo-1, dsDNA, nucleosomes, histones and ribosomal P protein were extracted from calf thymus by acetone method Natural protein extracted from tissue; AMA-M2 natural protein extracted from pig heart tissue. The specific method is as follows:

[0037] Calf thymus and pig heart were frozen and stored in -80°C ultra-low temperature freezer. First, ANA was extracted with phosphate buffered saline (PBS) and Tris-HCl buffer at 4°C, and precipitated with -20°C pre-cooled acetone Extract ANA into dry powder, do polyacrylamide gel electrophoresis (SDS-PAGE) and vertical transfer (Western blot) together with Sigma rabbit thymus acetone powder, transfer to nitrocellulose membrane, ID and IBT detect anti-ANA antibody, Secondly, different saturation concentrations of ammonium sulfate were used to precipitate pig ...

Embodiment 2

[0049] Embodiment 2 The usage method of kit of the present invention

[0050] Sample requirements: Take 2-5ml of blood from the vein, put it in a test tube and centrifuge the serum after coagulation, store the serum sample at 2-8°C, store the sample that cannot be tested within 24 hours at -20°C, severe hemolysis, floc or mold Altered serum samples will affect the test.

[0051] 1. Add sample

[0052] Dilute the sample to be tested with the sample diluent at a rate of 1:100, add 100ul to the reaction tank of the detection reagent prepared in Embodiment 1 above, and put it into an incubation shaker. React at 37°C for 30 minutes.

[0053] 2. Washing board

[0054] After the sample reaction, the reaction solution was discarded, and shaken and washed for 3×5min.

[0055] 3. Add enzyme-labeled secondary antibody

[0056] Alkaline phosphatase-labeled goat anti-human IgG antibody was diluted 10 times, and 100ul was added to each reaction tank, placed in an incubation shaker, and...

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Abstract

The invention discloses a kit used for detecting the relevant ANA spectrum of self-immune disease, and a preparation method thereof. The kit comprises a reaction card, an enzyme-linked reagent, a developer, a stopping solution and a concentrated washing solution; the reaction card consists of a carrier sheet and a nitrocellulose film or a nylon film which is fixed on the carrier sheet; the nitrocellulose film or the nylon film comprises 15 parallel detection lines which are wrapped respectively by nRNP / Sm, Sm, SS-A, Ro-52, SS-B, Scl-70, Jo-1, nucleosome, dsDNA, histone, ribosome P protein, AMA-M2, PM-Scl, CENP B, PCNA15 natural and recombinant antigen. The kit of the invention can exactly diagnose the self-immune disease, overcomes the shortages of the existing immunoblot diagnosis method on detecting the self-immune disease, and has the advantages of high sensitiveness, strong specificity, good exactness, simple and convenient operation, little consumption of the samples, and objective result judging, etc.

Description

technical field [0001] The present invention relates to a kit for diagnosing diseases, in particular to a kit for detecting autoimmune disease related anti-nucleic antibody (anti-nucleic antibody, ANA) spectrum, the present invention also relates to the preparation method and use of the diagnostic kit The method belongs to the field of biomedicine. Background technique [0002] Autoimmune diseases are diseases in which the immune system reacts to the components of its own body, causing damage. Common autoimmune diseases include: systemic lupus erythematosus, rheumatoid arthritis, scleroderma, hyperthyroidism, juvenile diabetes, idiopathic thrombocytopenic purpura, autoimmune hemolytic anemia, ulcerative colitis, and many skin disease, chronic liver disease, etc. [0003] Under normal circumstances, the immune system only responds to foreign substances that invade the body, such as bacteria, viruses, parasites, and transplants, and destroys or rejects these foreign substanc...

Claims

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Application Information

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IPC IPC(8): G01N33/558
Inventor 金鑫戴路
Owner BEIJING MOKOBIO LIFE SCI CO LTD
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