ELISpot技术检测猪抗口蹄疫γ干扰素方法的建立

doi:10.16431/j.cnki.1671-7236.2017.01.007

Abstract

Abstract:

The food and mouth disease virus (FMDV) vaccination is a key method in prevention and control of FMD, and the assay of antibodies to FMDV is always used to evaluate the immunity and preventive ability to FMD for its simplicity. However, the antibody titer is not always paralleled with the immunity to FMDV, for cellular immunity is also an important factor in FMDV response. So, we set up and optimized a ELISpot technique to detect specific interferon gamma(IFN-γ) to FMDV. Both the peptide pool and whole virus particle of FMDV(O/Mya98/XJ/2010) were used as stimulators, and the phytohaemagglutinin (PHA) was used as positive control. The final concentration of PBMC, whole virus and peptide pool and incubation time were optimized. The results showed that the optimized condition for ELISpot to detect specific IFN-γ to FMDV were as followed, more than 90% viability of fresh isolated or cryopreserved PBMC were seeded at the suitable concentration of 2×10⁵ cells/well, stimulated with whole virus particle containing 100 ng/mL 146 S or T cell epitope peptide pool consisting of 10 μg/mL each peptide or 20 μg/mL PHA and incubated for 16 hours. The foundation of ELISpot technique to detect specific IFN-γ to FMDV would provide a valuable means to evaluate the cellular immunity to FMDV of swine inoculated with FMD vaccine. The relevance of specific IFN-γ level to FMDV with 50% preventive dose (PD₅₀) of challenge test needed to be further investigated in the future.

Key words: ELISpot; foot and mouth disease; IFN-γ

CLC Number:

S852.65

LIU Jia, GAO Xi-xi, LI Wei, CHENG Shu-jing, LI Guang-yan, YU Han-xun, WANG Qin-fu. Establishment of ELISpot Technique for Detecting IFN-γ against Foot and Mouth Disease Virus[J]. , 2017, 44(1): 53-58.

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Establishment of ELISpot Technique for Detecting IFN-γ against Foot and Mouth Disease Virus

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