中国畜牧兽医2017，Vol.44Issue(1)：53-58,6.DOI:10.16431/j.cnki.1671-7236.2017.01.007
ELISpot技术检测猪抗口蹄疫γ干扰素方法的建立
Establishment of ELISpot Technique for Detecting IFN-γagainst Foot and Mouth Disease Virus
摘要
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 immuni-ty and preventive ability to FMD for its simplicity.However,the antibody titer is not always par-alleled 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 con-trol.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-γto 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 parti-cle 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-γto FMDV would provide a valuable means to evaluate the cellular immunity to FM-DV of swine inoculated with FMD vaccine.The relevance of specific IFN-γlevel to FMDV with 50% preventive dose (PD50 )of challenge test needed to be further investigated in the future.关键词
ELISpot/口蹄疫/γ干扰素Key words
ELISpot/foot and mouth disease/IFN-γ分类
农业科技引用本文复制引用
刘佳,高习习,李巍,程淑晶,李广焱,于汉勋,王钦富..ELISpot技术检测猪抗口蹄疫γ干扰素方法的建立[J].中国畜牧兽医,2017,44(1):53-58,6.基金项目
国家自然科学基金（81172777）；大连市科技计划基金 （）
