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Hybridoma cell strain capable of secreting CPIV3 antibody and ELISA kit

A technology of hybridoma cell lines and kits, applied in the field of hybridoma cell lines and ELISA kits

Active Publication Date: 2016-08-10
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no high-throughput detection method for goat parainfluenza virus (CPIV3) antibody at home and abroad. This invention aims to establish a blocking ELISA detection kit for detecting CPIV3 antibody, laying the foundation for clinical detection and related research.

Method used

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  • Hybridoma cell strain capable of secreting CPIV3 antibody and ELISA kit
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  • Hybridoma cell strain capable of secreting CPIV3 antibody and ELISA kit

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Experimental program
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preparation example Construction

[0020] 1. Antigen preparation: Infect MDBK with CPIV3 virus (JS2013) (Yang L, Li W, Mao L, et al. Analysis on the complete genome of novel caprine parainfluenza virus 3. Infect GenetEvol.2016,38:29-34.) Cells were inoculated for 4 days, and the virus was harvested after cell lesions appeared, and centrifuged at 10,000rpm for 30min to remove cell debris; the supernatant was ultracentrifuged at 40,000g for 2h, the supernatant was discarded, and the precipitate was dissolved overnight with an appropriate amount of PBS.

[0021] 2. Immunization of Balb / c mice: immunize Balb / c mice with multi-point subcutaneous injection of ultra-isolated virus, a total of 3 immunizations, with an interval of 2 weeks between each immunization, and use 100ul of ultra-isolated virus and the same amount of complete Freund's adjuvant for the first immunization Mixed immunization with 100ul ultra-isolated virus and the same amount of incomplete Freund's adjuvant for the last two times; blood was collecte...

Embodiment

[0050] 212 goat clinical serum samples were collected, and the CPIV3 antibody in the serum was detected by the blocking ELISA method and neutralization test established above, respectively. Compared with the results of blocking ELISA and neutralization experiment, the coincidence rate was 98.8%.

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Abstract

The invention belongs to the field of molecular biology, and relates to a hybridoma cell strain capable of secreting a CPIV3 antibody and an ELISA kit. A caprine parainfluenza virus 3 CPIV3 immunogen used in the invention is a CPIV3 JS2013 viral strain isolated clinically, and a hybridoma cell strain 2E6 is screened from an established hybridoma cell bank capable of secreting the CPIV3 antibody. The monoclonal antibody secreted by the hybridoma cell strain has the blocking effect, and is applied to the CPIV3 blocking ELISA kit. The CPIV3 blocking ELISA kit can specifically detect the antibody generated after the CPIV3 infection or caprine immunization. 212 caprine clinical serum samples are colleted, and the CPIV3 antibody in the serum is tested by the established blocking ELISA method and a neutralization test. Through the comparison between the result of the blocking ELISA method and the result of the neutralization test, the coincidence rate is 98.8%.

Description

technical field [0001] The invention belongs to the field of molecular biology and relates to a hybridoma cell line secreting CPIV3 antibody and an ELISA kit. Background technique [0002] Parainfluenza virus type 3 (PIV3) belongs to the Respiratory Virus genus of the Paramyxoviridae family and is a single-stranded negative-sense RNA virus with an envelope. HPIV3), bovine parainfluenza virus type 3 (BPIV3), etc. Members of this genus are important pathogens that cause respiratory diseases in humans and animals. At the end of 2013, our laboratory reported for the first time that a new type of PIV3 was found in sheep with respiratory diseases in Jiangsu and other places. The genetic evolution analysis was obviously different from HPIV3 and BPIV3, and it was named caprine parainfluenza virus type 3 (CPIV3). Epidemiological surveys show that CPIV3 is more common in flocks, causing greater losses due to disease. [0003] PIV3 has six structural proteins, namely N, P, M, F, HN ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/10G01N33/577C12R1/91
CPCC07K16/1027G01N33/577
Inventor 毛立李文良江杰元杨蕾蕾郝飞李基棕张纹纹周天赐
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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