Method for testing potency of duck hepatitis A bivalent live vaccine
A duck hepatitis A and detection method technology, which is applied in the field of duck hepatitis A bivalent live vaccine efficacy testing, can solve problems such as inaccurate vaccine potency results, achieve important use value and promotion prospects, and break through technical bottlenecks
Pending Publication Date: 2021-12-24
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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Problems solved by technology
[0004] The purpose of the present invention is to address the deficiencies in the prior art and provide a method for testing the efficacy of duck hepatitis A bivalent live vaccines, to overcome the mutual influence of the neutralization test between different serotypes of duck hepatitis A virus, resulting in vaccine failure. Technical issues with inaccurate titer results
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Embodiment
[0028] The method for testing the effectiveness of the duck hepatitis A bivalent live vaccine of the present invention will be described in detail below in conjunction with the examples.
[0029] 1. Antigen preparation and lyophilization
[0030] 1. Preparation of serotype 1 duck hepatitis A virus A66 strain antigen:
[0031] The DHAV-1A66 strain of Duck hepatitis virus was taken and sent to No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, and Chinese Microbial Strains on August 13, 2013. Preserved by the General Microbiology Center of the Preservation Management Committee, the preservation number is CGMCC No.8020), diluted 100-1000 times with sterile normal saline, inoculated with 7-9-day-old SPF chicken embryos, 0.2ml per embryo, and placed at 36-37°C to continue After incubation, the dead embryos were discarded within 24 hours, and the chicken embryo tissues (fetus, embryo membrane) that died within 24 to 9...
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The invention provides a method for testing the potency of a duck hepatitis A bivalent live vaccine, which comprises the following steps: inoculating mixed antigens containing serum type 1 duck hepatitis A and serum type 3 duck hepatitis A into 7-9-day-old SPF chick embryos, discarding the chick embryos dead within 24 hours, collecting all chick embryo tissues 24-120 hours after inoculation, carrying out total RNA extraction, carrying out RT-PCR detection by respectively utilizing DHAV-1 and DHAV-3 specific primers, carrying out agarose gel electrophoresis, then counting the PCR positive number, and calculating the chicken embryo half infection dose (EID50) of the two antigens, so as to avoid the mutual influence of the two viruses during the chicken embryo virus titer determination. The two duck hepatitis A viruses provided by the invention have good adaptability to inoculation proliferation of the SPF chick embryos, and are particularly suitable for the detection method provided by the invention.
Description
Technical field: [0001] The invention belongs to the technical field of vaccine production, in particular to a method for testing the efficacy of duck hepatitis A bivalent live vaccine. Background technique: [0002] Duck hepatitis A virus (DHAV) is the only member of the genus Avian Hepadnavirus in the small RNA family. After the virus infects ducklings less than 3 weeks old, the mortality rate reaches more than 90%, and it can occur all over the world. Duck hepatitis A was first reported in the United States in 1945, and the virus was successfully isolated for the first time in 1950. So far, DHAV has been divided into three serotypes, DHAV-1, DHAV-2, and DHAV-3, through the analysis of phylogenetic trees and relevant data such as serum neutralization tests, among which serotype 1 duck hepatitis A virus (DHAV-1) As a classic strain, serotype 2 duck hepatitis A virus (DHAV-2) has only been reported in Taiwan, China. [0003] As we all know, my country is a big duck breedi...
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IPC IPC(8): C12Q1/70C12Q1/686C12N7/00C12R1/93
CPCC12Q1/706C12Q1/686C12N7/00C12N2770/32421C12Q2565/125
Inventor 卢凤英张小飞吴凤瑶赵莎刘青涛沈丽雅
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES