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Homogeneous immunodetection kit for detecting anti-CCP antibody and application thereof

An immunodetection and anti-immune technology, applied in the field of immunoassay, which can solve the problems of low sensitivity, false positives, and narrow linear range.

Active Publication Date: 2019-08-23
CHEMCLIN DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the anti-cyclic citrullinated peptide antibody detection kits on the domestic and foreign markets mainly use the enzyme-linked immunosorbent assay (ELISA method), but the detection background value of this method is relatively low. High, low sensitivity, narrow linear range, easy to produce false positive experimental results; at the same time, for the detection of different subtypes of antibodies, corresponding enzyme-labeled secondary antibodies are required for detection, and the experimental operation is relatively cumbersome

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  • Homogeneous immunodetection kit for detecting anti-CCP antibody and application thereof
  • Homogeneous immunodetection kit for detecting anti-CCP antibody and application thereof
  • Homogeneous immunodetection kit for detecting anti-CCP antibody and application thereof

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preparation example Construction

[0106] In the present invention, there is no special limitation on the preparation method of polyclonal antibody, and it can be prepared by conventional preparation methods in the field. For example, the preparation method of the polyclonal antibody includes: immunizing animals with human immune complexes, obtaining the Animal serum of polyclonal antibodies; the animal serum is purified by affinity chromatography to obtain polyclonal antibodies that specifically recognize human immune complexes.

[0107] In some specific embodiments of the present invention, the preparation method of the polyclonal antibody comprises the following steps:

[0108] L1, immunize animals with the first group of human immune complexes, and collect animal serum after immunization;

[0109] L2, the animal serum is loaded onto the affinity chromatography column bound to the second group of human immune complexes, and after washing and eluting, the elution of polyclonal antibodies that specifically rec...

Embodiment 1

[0293] Example 1: Preparation of polyclonal antibodies that specifically recognize human immune complexes as anti-immune complex antibodies

[0294] (1) Inducing immune tolerance to human IgG in New Zealand white rabbits

[0295] Purchase 2.5 grams of human immunoglobulin for injection, concentration 50mg / mL, take 20mL for dialysis in normal saline, and change the dialysate 3 times during the period. The dialyzed human immunoglobulin was centrifuged at 100,000 g for 90 min, and the upper 1 / 3 liquid was taken to obtain monomeric human IgG, and its concentration was measured.

[0296] Eight male New Zealand white rabbits with a body weight of about 2.5 kg were taken, and each rabbit was injected with 10 mg of monomeric human IgG into the ear margin vein to induce immune tolerance of New Zealand white rabbits to human IgG.

[0297] (2) Preparation of immune complexes and immunization of rabbits with the immune complexes

[0298] With a 5mL syringe, draw 2mL of Alfred's solution...

Embodiment 2

[0317] Example 2: Preparation of monoclonal antibodies that specifically recognize human immune complexes as anti-immune complex antibodies

[0318] (1) Induce mouse immune tolerance to human IgG

[0319] Purchase 2.5 grams of human immunoglobulin for injection, concentration 50mg / mL, take 20mL for dialysis in normal saline, and change the dialysate 3 times during the period. The dialyzed human immunoglobulin was centrifuged at 100,000 g for 90 min, and the upper 1 / 3 liquid was taken to obtain monomeric human IgG, and its concentration was measured.

[0320] Six male Balb / c mice aged 6-8 weeks were taken, and each mouse was injected with 2 mg monomeric human IgG through the tail vein to induce immune tolerance to human IgG in mice, and the formal immunization was performed one week later.

[0321] (2) Preparation of immune complexes and immunization of mice with the immune complexes

[0322] Take 3 mice, enucleate the eyeballs, collect the whole blood into a 15mL centrifuge ...

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Abstract

The invention relates to a homogeneous immunodetection kit for detecting an anti-CCP antibody. The kit adopts high sensitivity and specificity of citrullinated bovine serum albumin as an antigen, andis prepared by combining an anti-immune complex antibody which can distinguish antibodies, non-specific antibodies and free specific antibodies that do not bind with antigens and a photo-stimulated chemiluminescence technology. The kit can omit washing processes in traditional indirect methods without being disturbed by non-specific antibodies during detection; and the kit is high in specificity,good in sensitivity, wide in linearity range, fast in reaction speed and convenient in operation, and can realize fully automated high throughput testing.

Description

technical field [0001] The invention belongs to the technical field of immune analysis, and in particular relates to a homogeneous immunoassay kit for detecting anti-CCP antibodies and a preparation method and application method thereof. Background technique [0002] Anticyclic citrullinated peptide antibodies (Anticyclic citrullinated peptide antibodies, anti-CCP) antibody is a new type of serum marker, clinical practice shows that the titer of anti-CCP antibody is usually related to the activity of the disease, 70-80% of patients in the Anti-CCP antibodies appear in serum and synovial fluid very early in the disease, and this antibody has high specificity and certain prognostic value. [0003] At present, the anti-cyclic citrullinated peptide antibody detection kits on the domestic and foreign markets mainly use the enzyme-linked immunosorbent assay (ELISA), but this method has a high background value, low sensitivity, narrow linear range, and is prone to false positives. ...

Claims

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Application Information

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IPC IPC(8): G01N33/558G01N33/532
CPCG01N33/558G01N33/532
Inventor 饶星廖智星刘宇卉李临其他发明人请求不公开姓名
Owner CHEMCLIN DIAGNOSTICS CO LTD
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