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Brucella antibody detecting test strip

A technology for detecting Brucella and antibodies, which is applied in the direction of measuring devices, instruments, scientific instruments, etc.

Inactive Publication Date: 2017-05-31
CHINA INST OF VETERINARY DRUG CONTROL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Chen Chuangfu from Shihezi University in Xinjiang, etc. (application number: CN201210254837.X) also used Staphylococcus aureus protein A (SPA) as the gold-labeled antibody, but the Brucella-specific proteins (OMP31 and BP26) expressed in vitro replaced Brucella As a detection antigen, LPS cannot fundamentally solve the problem of sensitivity to different animal diagnoses
Hangzhou Dean Technology Co., Ltd. Zhang Mingzhou et al. (application number: CN201310033074.0) used Brucella bacterium antigen as the quality control line to coat the antigen, and then used an anti-erythrocyte monoclonal antibody as the monoclonal antibody for competition. Adsorbed dry colloidal gold protein (flow band), established a sandwich method to detect the bovine Brucella antigen detection test card, because Brucella and some Gram-negative bacteria, especially Yersinia O9 and Escherichia coli Bacillus O157 has strong bacterial antigen similarity, and the patent uses Brucella bacterial antigen as the adsorption antigen of colloidal gold particles, theoretically, the detection results will have certain non-specificity

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] ——Preparation of test strips

[0041] 1. Preparation of test strip LPS antigen

[0042] The Brucella vaccine S2 strain was selected as the seed for antigen production. After S2 was cultured with TSA, the extracted LPS was purified and quantified and used to fix the nitrocellulose membrane on the colloidal gold test strip at the position corresponding to the detection line.

[0043] (1) Establish quality standards for Brucella S2 pigs for LPS antigen preparation.

[0044] 1) Colony morphology The edge of the colony is neat, round, dewdrop-shaped, irradiated with oblique light, and observing against the light with a slight blue opalescence.

[0045] 2) The staining form is coccobacteria, scattered individually, without forming spores and capsules. The size is between 0.3 and 0.6 μm. Gram stain is negative, Korotk stain is red.

[0046] 3) It should be carried out purely according to the appendix of the current "Chinese Veterinary Pharmacopoeia".

[0047] 4) Specificity

[0048] ① Se...

Embodiment 2

[0071] ——The source of the main raw materials and the inspection methods of the test strips for the detection of Brucella antibodies

[0072] 1. Flag monoclonal antibodies were purchased from Sigma. For different batches of Flag monoclonal antibodies, the following tests should be carried out before use:

[0073] (1) Properties are colorless and clear liquid.

[0074] (2) The product analysis report provided by the supplier checks the product analysis report provided by the supplier to verify whether the protein content, molar ratio, specificity and other parameters are within the specified range.

[0075] (3) The sterility test is carried out in accordance with the appendix of the current "Chinese Veterinary Pharmacopoeia" (Chinese Veterinary Pharmacopoeia Committee, Veterinary Pharmacopoeia of the People's Republic of China, 2010 Edition Three, China Agriculture Press, 2011, hereinafter referred to as "Chinese Veterinary Pharmacopoeia") Inspection, there should be no bacterial growt...

Embodiment 3

[0087] ——Competitive ELISA method for determination of monoclonal antibody (3E4) ascites titer

[0088] 1. Ascites dilution: Dilute the prepared ascites with 1×PBS solution at 1:10000, 1:15000, 1:20000, 1:25000, 1:30000, 1:35000 and 1:40000.

[0089] 2. cELISA assay Take the antigen-coated plate (according to the number of samples, it can be disassembled for separate use), wash the plate once with 1×washing solution 300μl / well, and discard the washing solution. The diluted positive control serum and negative control serum were added to the ELISA plate, 50μl / well, and the positive and negative control sera were repeated twice. After adding the sample, add the diluted monoclonal antibody, 50μl per well, shake and mix for 5min. After incubating at 37°C for 30 minutes, take out the reaction plate, discard the reaction solution, add 300 μl 1× washing solution to each well, wash 3 times, and spin dry. After diluting the HRP-labeled goat anti-mouse IgG antibody 1:100 with the correspond...

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Abstract

The invention relates to a Brucella antibody detecting test strip. The Brucella antibody detecting test strip has the following significance: (1) Brucellosis diagnostic methods are enriched; (2) one novel method which is simple and fast and can implement rapid field detection on various animals is provided for Brucellosis diagnosis in China; (3) the test strip overcomes the defect that the conventional Brucellosis diagnostic methods cannot eliminate interference of enterocolitis Yersinia O9 and escherichia coli O157 antiserum in Brucella antibodies, and improves the Brucellosis detecting specificity.

Description

Technical field [0001] The invention relates to a Brucella antibody detection test strip, which belongs to the technical field of biological product detection. [0002] technical background [0003] Brucellosis (Brucellosis, referred to as brucellosis) is a zoonotic disease characterized by abortion and fever caused by Brucella or Brucella, which seriously threatens humans and many animals Life and health. This disease not only has serious harm to the reproduction and production performance of animals, but more importantly, it is often difficult to cure people infected with Brucella, which causes serious public health problems. Therefore, in countries where Brucella is endemic, the elimination of brucellosis has always been one of the most important goals in public health plans. [0004] Brucellosis has a long history in the world. Humans have gradually deepened their research understanding of the disease, and the diagnostic level has been continuously improved. With the developmen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577
CPCG01N33/577
Inventor 丁家波冯宇朱良全蒋卉彭小薇王芳王楠毛开荣徐磊
Owner CHINA INST OF VETERINARY DRUG CONTROL
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