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O-type foot-and-mouth disease virus antibody chemiluminescence detection kit

A technology for chemiluminescence detection and foot-and-mouth disease virus, which is applied in chemiluminescence/bioluminescence, analysis through chemical reactions of materials, and measurement devices, etc., to achieve strong specificity, low carcinogenicity and harm, and stable markers

Inactive Publication Date: 2016-12-14
洛阳现代生物技术研究院有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Traditional FMD diagnosis and quarantine methods mainly include serological methods such as complement fixation test, agar diffusion test, neutralization test, indirect hemagglutination test, enzyme-linked immunosorbent assay, etc. The antibodies used are prepared from the antiserum of immunized animals. Antiserum is actually an antibody complex, which is prone to strong non-specificity when used as a diagnostic reagent

Method used

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  • O-type foot-and-mouth disease virus antibody chemiluminescence detection kit
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  • O-type foot-and-mouth disease virus antibody chemiluminescence detection kit

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Experimental program
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Effect test

Embodiment 1

[0037] A chemiluminescent detection kit for O-type foot-and-mouth disease virus antibody, comprising polystyrene plate coated with antigen, monoclonal antibody, standard, luminescent substrate A solution and luminescent substrate B solution;

[0038] The coated antigen polystyrene plate is an opaque polystyrene plate coated with O-type foot-and-mouth disease virus RE2 recombinant protein. The volume ratio of the buffer solution is 1:5000, and the coated antigen is diluted; the diluted coated antigen is added to a 48-well or 96-well opaque white microwell plate with a sample volume of 100 μl / well. Coating at 4°C for 20 hours, washing with washing solution 2-3 times, then adding 150 µl / well of blocking solution, blocking at 4°C for 16 hours, then throwing off the liquid, drying at 37°C for 3 hours, and sealing in bags;

[0039] The monoclonal antibody is the O-type foot-and-mouth disease virus monoclonal antibody labeled with horseradish peroxidase, and the working concentration...

Embodiment 2

[0060] Preparation of a kind of O-type foot-and-mouth disease virus antibody chemiluminescent detection kit:

[0061] 1. The foot-and-mouth disease O-type recombinant protein was used as an antigen to immunize Balb / c mice, and the optimal antigen coating concentration (2.5mg / L) was determined by square array titration indirect ELISA method to detect the mouse serum, and the titer reached 1:4.096 ×10 4 , the splenocytes were fused with Sp2 / 0 myeloma cells to prepare FMD type O monoclonal antibody. Positive hybridoma cell lines were screened by limited dilution method and established indirect ELISA method, and finally the specific hybridoma cell line that can stably secrete monoclonal antibody was obtained from the cell line with the highest positive value after 4 times of subcloning screening . The hybridoma cells have good stability through the in vitro passage test, continuous cryopreservation recovery test, and ascites continuous passage test; after the specific type detec...

experiment example

[0070] 1. Specificity test

[0071] The positive sera of FMD A, Asia-1, swine fever, porcine reproductive and respiratory syndrome virus and Escherichia coli were detected with 3 batches of trial-produced FMDV antibody chemiluminescent detection kits.

[0072]

[0073] 2. Sensitivity test

[0074] Dilute the sensitive positive quality control serum at 1:16, 1:32, 1:64, 1:128, 1:256, 1:512, 1:1024, 1:2048, from the 3 batches of test kits 1 kit was randomly selected from each batch for detection to determine the sensitivity of the O-FMDV antibody chemiluminescence detection kit.

[0075]

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Abstract

The invention relates to an O-type foot-and-mouth disease virus antibody chemiluminescence detection kit. The kit includes an polystyrene board coated by antigen, a monoclonal antibody, a standard substance, a luminescence substrate solution A and a luminescence substrate solution B. The polystyrene board coated by antigen is an opaque polystyrene plate coated by an O-type foot-and-mouth disease virus RE2 recombinant protein; the monoclonal antibody is a horseradish peroxidase labeled O-type foot-and-mouth virus monoclonal antibody; the luminescence substrate solution A comprises a Lumino, hydroxycoumarin, gallic acid, a Tris-Hcl buffer and water; the standard substance is foot-and-mouth virus O-type antibody respectively diluted buy a calibration diluent, wherein the dilution is 0NU / mL, 2NU / mL, 5NU / mL, 10NU / mL, 30NU / mL and 60NU / mL respectively. The kit of the invention can be used for detecting O-type foot-and-mouth disease virus antibody, has the advantages of high sensitivity and wide detection range, and realizes the quantitative detection of foot-and-mouth disease antibody.

Description

technical field [0001] The invention relates to a kit, in particular to a chemiluminescent detection kit for O-type foot-and-mouth disease virus antibody. Background technique [0002] Foot-and-mouth disease (FMD) is an acute, febrile, highly contagious infectious disease of cattle, pigs, sheep and other cloven-hoofed animals caused by foot-and-mouth disease virus (FMDV). Water scars appear on the mouth, hooves, nipples, etc., and ulcers form after the water scars rupture. The outbreak and prevalence of the disease often cause huge losses to animal husbandry production and economic development, mainly due to the increase in the death rate of sick animals, the decline in production performance, and the loss of seed value. More importantly, any country and region does not cooperate with Trade between countries with foot-and-mouth disease has seriously affected the development of economic trade and animal husbandry, and caused related political, economic and social problems, s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/531G01N33/543G01N21/76
CPCG01N33/56983G01N21/76G01N33/531G01N33/54306
Inventor 李秀梅耿玉静李权伟刘玉宝张鹏翼杨志
Owner 洛阳现代生物技术研究院有限公司
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