Cyclic citrullinated peptide double-microsphere conjugate as well as preparation method and application thereof

A technology of cyclic citrulline peptides and conjugates, applied in the biological field, can solve the problems of low production efficiency, high production cost, and cumbersome preparation process, and achieve the effects of simple preparation process, improved sensitivity, and good stability

Pending Publication Date: 2021-03-16
海丰生物科技(北京)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It has the defects of cumbersome preparation process, low preparation efficiency and high manufacturing cost

Method used

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  • Cyclic citrullinated peptide double-microsphere conjugate as well as preparation method and application thereof
  • Cyclic citrullinated peptide double-microsphere conjugate as well as preparation method and application thereof
  • Cyclic citrullinated peptide double-microsphere conjugate as well as preparation method and application thereof

Examples

Experimental program
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Embodiment 1

[0021] Example 1, the preparation method of the cyclic citrullinated peptide double microsphere conjugate of the present invention

[0022] The preparation method of the cyclic citrulline peptide double microsphere conjugate of this embodiment comprises the following steps:

[0023] Step 1. Activation of carboxyl latex microspheres

[0024] Add 1mL of 10% polystyrene latex microspheres with a diameter of 120nm to 20mL of 0.05M MES buffer at pH 6.5, then add 5mL of carbodiimide aqueous solution with a concentration of 1mg / mL, stir and activate at room temperature for 30min to obtain activated Polystyrene latex microspheres, wherein the particle size range of the latex microspheres is 80-200nm;

[0025] Step 2. Coupling of carboxyl microspheres and cyclic citrullinated peptides

[0026] Dilute 5mg of cyclic citrullinated peptide with 50ml of 0.05M pH8.0 MOPS buffer, mix well, pour into the above-mentioned activated polystyrene latex microsphere emulsion, and stir at room tempe...

Embodiment 2

[0038] Example 2, Anti-cyclic citrullinated peptide antibody latex immunoturbidimetric detection kit

[0039] Take the cyclic citrulline peptide double microsphere conjugate prepared in Example 1 of the present invention as the second reagent component of the kit, and match the first reagent component mainly composed of buffer (Table 2) and calibrator components, and combine A detection kit can be used for quantitative detection of cyclic citrullinated peptide antibody.

Embodiment 3

[0040] Example 3. Preparation of products for diagnosing rheumatoid arthritis and its application to the detection of patients with rheumatoid arthritis

[0041] Serum samples from patients with rheumatoid arthritis were taken, and the anti-cyclic citrullinated peptide antibody latex immunoturbidimetric detection kit prepared in Example 2 of the present invention was used to detect the level of anti-cyclic citrullinated peptide antibodies in the patient samples for use in Auxiliary diagnosis of rheumatoid arthritis.

[0042] 1. Preparation of reaction reagents

[0043] The preparation of reagent one (R1) is shown in Table 2, the composition formula of R1 reagent.

[0044] Table 2

[0045]

[0046]

[0047] Preparation of reagent two (R2):

[0048]The amino microsphere-cyclic citrullinated peptide-carboxyl microsphere conjugate was prepared according to Example 1, and the final 0.2% conjugate was reagent 2 (R2).

[0049] 2. The machine test of amino microsphere-cyclic...

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Abstract

The invention discloses a cyclic citrullinated peptide double-microsphere conjugate as well as a preparation method and application thereof, and the method comprises the step of connecting carboxyl latex microspheres and amino microspheres with cyclic citrullinated peptide to form an amino microsphere-cyclic citrullinated peptide-carboxyl microsphere conjugate. According to the invention, two microspheres with approximate particle sizes are utilized, carboxyl microspheres are firstly used, and then amino microspheres are used for coupling with CPP antigens, so that the sensitivity of the detection reagent is improved, and meanwhile, a relatively good detection linear range can be kept; and a reagent prepared by the method provided by the invention has good stability, and the preparation process of the reagent is simple.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a cyclic citrullinated peptide double microsphere conjugate and its preparation method and application. Background technique [0002] RA is the most common joint inflammatory disease. The incidence rate of adults in developed countries is 1-2%. The etiology is unknown. It can occur at any age, but it is common in women aged 35-55. RA is a chronic disease and develops in stages Characterized by alternating with remission. [0003] Anti-CCP antibody was found to be useful in the early diagnosis of rheumatoid arthritis, and has certain predictive value for joint invasion and radiological damage in RA patients. Anti-CCP antibodies can be detected in RA patients before overt joint damage occurs. A prospective cohort study showed that among anti-CCP positive patients without obvious clinical symptoms of RA, about 93% of the patients would develop RA, indicating that the antibody has a ve...

Claims

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Application Information

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IPC IPC(8): B01J13/14C07K16/18G01N33/68G01N33/546G01N33/543
CPCB01J13/14C07K16/18G01N33/6893G01N33/54313G01N2800/102
Inventor 石怀兴温志国
Owner 海丰生物科技(北京)有限公司
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