ELISPOT,MHC肽五聚体和ICCD三种检测特异性细胞免疫应答方法的比较研究

Comparison of ELISPOT,MHC pentamer and ICCD methods for their capacity to detect the specific cell-mediated immune response

目的比较三种特异性细胞免疫应答的检测方法,寻求一种简便、重复性好、易于质控的评价疫苗细胞免疫的检测方法。方法采用HIVDNA疫苗肌内注射BALB/c小鼠,第6周用艾滋病重组MVA痘苗病毒腹腔注射加强免疫,于第10天制备脾脏细胞悬液,并用酶联免疫斑点法(Enzyme-Linked Immune Spot,ELISPOT)、MHC肽五聚体法和胞内细胞因子染色分析法(Intra-cellular cytokinedetection,ICCD)分别检测细胞免疫应答。结果ELISPOT、MHC肽五聚体染色以及胞内细胞因子染色分析法检测疫苗组细胞免疫应答的阳性率分别为5/5、4/5和3/5;而且ELISPOT和MHC肽五聚体染色法之间有一定相关性(r=0.647),而ELISPOT和胞内因子染色法之间以及MHC肽五聚体染色和胞内因子染色之间无相关性。结论ELISPOT试验操作相对简单,灵敏度高、重复性好,是评价疫苗细胞免疫的有效方法。

Objective To seek more convenient, repeatable and sensitive and easier method for evaluating the cellular immune effect of vaccine by comparing 3 different methods. Methods HIV DNA vaccine was administered to BALB/c mice via muscle injection, which was boosted with recombinant MVA containing HIV target gene via intraperitoneal injection at the 6th week after immunization with HIV vaccine, Splenic cells were prepared on the 10th day after immunization with MVA. Enzyme-Inked immune spot test （ELISPOT）, MHC-Pentamer and intra-cellular cytokine detection （ICCD） were used to measure the effect of cellular immune response. Results The cellular immune response in vaccined group was 5/5, 4/5 and 3/5 by using ELISPOT, MHC-Pentamer and ICCD respectively. The results of ELISPOT and MHC-Pentamer were correlated at medium level（ r = 0.647） . However, no relationship between ELISPOT and ICCD and between MHC-Pentamer and ICCD were indicated in this study. Conclusion ELISPOT would be used as quality control method for evaluation of vaccine.