Method for detecting valence of antibody

A detection method and technology of antibody titer, applied in chemical instruments and methods, biological testing, material testing products, etc., can solve the problems of immature detection methods and long detection time, and achieve good reactivity, good linearity, and reaction conditions. mild effect

Inactive Publication Date: 2013-12-11
SOUTHERN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Other detection methods are mostly in the initial stage of exploration and research. Due to the limitations of detection equipment, reagent prices, etc., the detection methods are still immature, and the factors that affect detection and diagnostic performance still need to be further improved.
However, the detection time of the existing ELISA method is still too long, generally it takes 2 hours to obtain the result

Method used

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  • Method for detecting valence of antibody

Examples

Experimental program
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preparation example Construction

[0040] Preparation of streptavidin-labeled polystyrene microspheres

[0041] 1) Take 0.1mL carboxylated latex microspheres, add 0.9mL ethanesulfonic acid buffer (MES) and mix well, then add water-soluble carbodiimide cross-linking agent (EDC) and N-hydroxysuccinimide (NHS) Activation, EDC concentration 1.0-1.5g / L, ratio of EDC to NHS 1:1-1:2, gentle stirring at room temperature, time controlled at 15-30min;

[0042] 2) Wash by centrifugation, centrifuge at 12000-15000g for 10-30min, wash the pellet three times with 0.1mol / L phosphate buffer (PBS), remove the supernatant, resuspend the pellet in 0.1mol / L PBS, oscillate, and sonicate Promptly obtain the activated latex microsphere;

[0043] 3) Dissolve 2 mg of streptavidin in PBS buffer with pH 6.5-7.8, add it to 1 mL of activated latex microsphere solution, stir gently at room temperature, and react for 2-4 hours;

[0044] 4) After the reaction time is reached, centrifuge at 12000-15000g for 10-30min, precipitate the micros...

Embodiment 1

[0048] 1) Dissolving CCP polypeptide: the prepared CCP polypeptide was dissolved in 0.15M pH9.0 phosphate buffer and stored at 4°C;

[0049] 2) Dissolving esterified biotin: dissolve esterified biotin with dimethyl sulfoxide (DMSO);

[0050] 3) Modification: quickly mix the dissolved CCP polypeptide solution with the dissolved esterified biotin solution, the ratio of biotin to CCP is 10:1, stir and incubate at room temperature for 4 hours;

[0051] 4) Remove excess biotin with a desalting column;

[0052] 5) Elute the sample with 0.05M PBS buffer, pH7.6.

[0053]Collect the eluate and evaluate the labeling result according to the UV absorption spectrum: According to the comparison of the UV absorption spectrum between the biotin-labeled CCP product and the unbiotin-labeled CCP at the same concentration (1mg / mL), the biotin-CCP between 210nm-230nm Increased absorbance values, such as image 3 . ELISA method was used to identify the effect of biotin-labeled CCP: the prepared...

Embodiment 2

[0056] 1) Dissolving CCP polypeptide: the prepared CCP polypeptide was dissolved in 0.2M pH9.6 phosphate buffer and stored at 4°C;

[0057] 2) Dissolving esterified biotin: dissolve esterified biotin with dimethyl sulfoxide (DMSO);

[0058] 3) Modification: quickly mix the dissolved CCP polypeptide solution with the dissolved esterified biotin solution, and the ratio of biotin to CCP is 10:1. Stir and incubate at room temperature for 4 h;

[0059] 4) Remove excess biotin with a desalting column;

[0060] 5) Elute the sample with 0.1M PBS buffer, pH7.2.

[0061] Collect the eluate and evaluate the labeling result according to the UV absorption spectrum: According to the comparison of the UV absorption spectrum between the biotin-labeled CCP product and the unbiotin-labeled CCP at the same concentration (1mg / mL), the biotin-CCP between 200nm-230nm Increased absorbance values, such as Figure 5 .

[0062] ELISA method was used to identify the effect of biotin-labeled CCP: th...

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Abstract

The invention discloses a method for detecting the valence of an antibody. The method comprises the following steps: 1), coupling an antigen on a microsphere to obtain a modified microsphere; 2), mixing and reacting the modified microsphere with an antibody solution; 3), measuring the turbidity of the reaction liquid, so as to determine the valence of the antibody. According to the method provided by the invention, CCP is modified on the surface of a polystyrene microsphere by adopting a vitamin H-streptavidin system, so as to prepare a granular CCP antigen successfully; in case that the granular CCP antigen prepared by adopting the method is used for detecting the valence of anti-CCP antibody in blood serum, the linearity of the detecting result is good, and the detecting process can be completed by 20 minutes, so that the method can be used for acquiring the detecting result more quickly as compared with the conventional ELISA detecting method consuming 2-3 hours.

Description

technical field [0001] The invention relates to a method for detecting antibody titer, in particular to a method for detecting antibody titer by using a modified solid phase carrier. Background technique [0002] Antibody titer is an important indicator in the fields of disease diagnosis, specific immunoglobulin preparation and vaccine evaluation, and its detection has very practical significance. Among the numerous antibody titer determination techniques, ELISA is the most widely used method because of its simple operation and fast speed. [0003] Rheumatoid arthritis (RA) is a chronic multisystemic inflammatory autoimmune disease mainly involving peripheral joints, characterized clinically by chronic inflammatory lesions in multiple peripheral joints, which can cause permanent The deformity of the joints and the high disability rate are called "undead cancer". Therefore, early diagnosis and early treatment can delay the progression of the disease, reduce the disability r...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68C07K17/08
Inventor 裘宇容付琳姜云飞
Owner SOUTHERN MEDICAL UNIVERSITY
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