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A-type H1 subtype influenza virus antibody blocking ELISA kit and applications thereof

A technology of influenza virus and kit, which is applied in the field of animal virology and immunology detection, can solve the problems of short detection time and achieve the effect of short detection time, simple operation and strong specificity

Inactive Publication Date: 2014-02-19
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The main purpose of the present invention is to overcome the defect of prior art, provide a kind of type A H1 subtype influenza virus antibody blocking ELISA detection kit, have strong specificity, high sensitivity, detection time is short, solve the problem of influenza A H1 subtype Problems with Influenza Virus Antibody Testing

Method used

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  • A-type H1 subtype influenza virus antibody blocking ELISA kit and applications thereof
  • A-type H1 subtype influenza virus antibody blocking ELISA kit and applications thereof
  • A-type H1 subtype influenza virus antibody blocking ELISA kit and applications thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0043] A type A H1 subtype influenza virus antibody blocking ELISA detection kit, the kit consists of the following materials:

[0044] a) A H1 subtype influenza virus A-Influ / JML-F9, which was sent to the China Center for Type Culture Collection (CCTCC) in Wuhan University, Wuhan, Hubei Province on January 27, 2011 for preservation, The deposit number is CCTCC NO: V201105. (Patent has been applied for, patent acceptance number: 201110036167.X, purchased at China Center for Type Culture Collection, please see the list of genetic resources)

[0045] b) A highly specific monoclonal antibody against the hemagglutinin (HA) protein of influenza A H1 subtype virus. The hybridoma cell line secreting the monoclonal antibody was sent to Wuhan, Hubei Province on January 27, 2011 Preserved at the China Center for Type Culture Collection (CCTCC) within Wuhan University, the preservation number is CCTCC NO: V201106 (patent applied for, patent acceptance number: 201110036167.X, purchased a...

Embodiment 2

[0087] A kind of H1 subtype influenza virus antibody blocking ELISA kit is used in the application of type A H1 subtype influenza virus antibody detection, and its steps are:

[0088] 1. Preparation of core reagents:

[0089] Coating solution (25mmol / L carbonate buffer): Na 2 CO 3 1.59g,NaHCO 3 2.93g, with ddH 2 O was adjusted to 1000mL (pH9.6).

[0090] 10 times washing liquid: NaCl 80g, KCl 2g, NaCl 2 HPO 4 12H 2 O 29g, KH 2 PO 4 2g, Tween-20 5mL, with ddH 2 O was adjusted to 1000mL (pH=7.4).

[0091] Blocking solution: 5g skimmed milk dissolved in 100mL washing solution.

[0092] Substrate solution: Divided into substrate solution A and substrate solution B. The specific composition is as follows:

[0093] Substrate solution A: 0.006% (volume ratio) concentration of H 2 o 2 buffer.

[0094] Substrate solution B: take Na 2 HPO 4 12H 2 O14.2g, citric acid 10.5g, with ddH 2 O is fixed to 500mM and made into 0.1mL phosphate citrate buffer solution (pH=5.0...

Embodiment 3

[0118] Sensitivity Test and Specificity Test of Blocking ELISA Detection Method for Influenza A H1 Subtype Antibody

[0119] 1. Sensitivity test of A / H1 subtype influenza virus antibody blocking ELISA detection method

[0120] Through the sensitivity test of the present invention, it is known that the HI titer of the positive serum is 1:2560. After ten-fold dilution of the original concentration, the serial two-fold ratio dilution shows the results as shown in the figure below. The sensitivity of this method can reach the HI titer of 2. 1(Yin Zhen et al. Animal Virology (Second Edition) [M]. Beijing: Science Press, 1997). see attached results figure 2 .

[0121] 2. Specificity test of ELISA detection method for antibody blocking of A / H1 subtype influenza virus

[0122] (1) Classical swine influenza H1 subtype (GD strain), classic swine influenza H1 subtype (TJ strain), swine influenza H3 subtype, swine fever, Japanese encephalitis, foot-and-mouth disease, pseudorabies, sma...

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Abstract

The invention discloses an A-type H1 subtype influenza virus antibody blocking ELISA kit. The kit comprises: a) an A-type H1 subtype influenza virus A-Influ / JML-F9; b) a monoclonal antibody with strong specificity of an A-type H1 subtype influenza virus-resistant hemagglutinin protein; c) an antibody blocking ELISA core kit prepared from the monoclonal antibody of the A-type H1 subtype influenza virus-resistant hemagglutinin protein; and d) a sample diluting liquid, 10 times of a washing liquid, a substrate liquid A, a substrate liquid B, a reaction stop solution, a positive control sample and a negative control sample in the kit. The invention also discloses applications of the A-type H1 subtype influenza virus antibody blocking ELISA kit in A-type H1 subtype influenza virus antibody detection. The kit has strong specificity, high sensitivity, short detection time, easy and practical operation, and no need of being operated by a professional person. The kit is good in stability and long in retention period, and solves a problem of the A-type H1 subtype influenza virus antibody detection.

Description

technical field [0001] The invention relates to the technical field of animal virology and immunology detection. Specifically, the present invention relates to a kind of A H1 subtype influenza virus antibody blocking ELISA detection kit, also relates to a kind of A H1 subtype influenza virus antibody blocking ELISA detection kit in swine population The use of influenza virus antibody detection. Background technique [0002] In March 2009, a large number of influenza-like cases and severe respiratory infections occurred in parts of Mexico. On April 15 and April 17, the US CDC detected the same new type of HA in two clinical samples with independent backgrounds. A subtype of influenza virus, the A(H1N1) virus, was subsequently identified as the causative agent of the influenza-like illness in March. In the following less than 6 weeks, the epidemic broke out in countries and regions other than the United States, Mexico, and Canada. In June 2009, the World Health Organization...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577C07K16/10
CPCG01N33/56983C07K16/1018G01N33/531
Inventor 金梅林李淑云王灿赵刚郭学波
Owner HUAZHONG AGRI UNIV
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