Detection method and kit for quantitatively detecting anti-cyclic guanidine polypeptide antibody
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A polypeptide antibody and detection method technology, applied in the field of biomedical detection, to achieve the effects of reducing detection costs, facilitating market promotion, and high sensitivity
Pending Publication Date: 2020-05-29
叶卫玲
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Embodiment 1
[0050] Embodiment 1 - the reagent of detection method of the present invention
[0056] Example 2——Preparation of CCP-carrier markers
[0057] The CCP-carrier labeling method and steps of the present invention are as follows:
[0058] 1. Prepare 100mM MES buffer solution, pH=6.0, prepare 10mM Tris buffer solution, pH=7.5, and set aside;
[0059] 2. Take 100mg BSA and dissolve it with MES buffer in step 1, the final concentration is 10mg / ml;
[0060] 3. Take 1 mg of CCP, add it to the solution in step 2, and stir at room temperature at low speed;
[0061] 4. Weigh 1 mg of EDC (carbodiimide), add it to the solution in step 3, stir at a low speed, and react at room temperature for 1 hour;
[0062] 5. Dialyze the reaction solution in step 4 through Tris buffer in step 1 for 4 times, and store at 2-8°C.
Embodiment 3
[0063] Example 3——Preparation of anti-CCP antibody
[0064] 1. Take 1mg of CCP, dissolve it in 20mM PBS, pH=7.5 buffer solution, the final concentration is 1mg / ml;
[0065] 2. Take the CCP solution in step 1 and Freund's incomplete adjuvant to prepare an emulsion to obtain an immune emulsion;
[0066] 3. Carry out rabbit (sheep) immunization with the immunization preparation in step 2, and the immunization cycle is about 120 days;
[0067] 4. Take the rabbit (sheep) immune serum and purify it by agarose affinity chromatography to obtain the rabbit (sheep) anti-CCP antibody.
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Abstract
The invention relates to a detection method and a kit for quantitatively detecting an anti-cyclic guanidine polypeptide antibody. The method for enhancing competitive turbidimetry by latex is adopted,An anti-cyclic citrulline polypeptide antibody in a to-be-detected sample and latex microspheres coated with antibodies in a reagent compete to be combined with cyclic guanidine acid polypeptide in the reagent, an absorbance value of the reagent is measured through a 500-700 nm wavelength spectrophotometric method, and a concentration value of the anti-cyclic guanidine acid antibody in the to-be-detected sample is calculated according to the attenuation amount of the absorbance value. The method is advantaged in that a linear range of the anti-cyclic guanidine acid antibody measured by the method can reach 7-500U/L, moreover, the kit is not interfered by rheumatoid arthritis and the like, and detection performance of the reagent can reach consistency of ELISA and electrochemical luminescence immunoassay. The detection kit of the method is high in sensitivity, good in specificity and suitable for a clinical chemical analyzer, moreover, detection cost is greatly reduced.
Description
technical field [0001] The invention relates to a detection method and a kit for quantitative detection of anti-cyclic guanidine polypeptide antibody. It belongs to the field of biomedical detection. Background technique [0002] Anticyclic peptide containing citrullin antibody (anticyclic peptide containing citrullin) is an autoantibody with synthetic cyclic citrullinated polypeptide (CCP) as its antigen. The detection of anti-CCP antibody is highly effective in the diagnosis of rheumatoid arthritis (RA). Specific, and can be used for early diagnosis of RA. It is generally believed that the sensitivity of anti-CCP antibody to the diagnosis of RA is 50%-78%, the specificity is 96%, and the positive rate of RA in early patients can reach 80%. In addition, anti-CCP antibody is not only a diagnostic indicator for the early diagnosis of RA, but also a sensitive indicator for distinguishing invasive and non-invasive RA. Antibody-positive patients tend to develop more severe jo...
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