结核分枝杆菌rCFP10—ESAT6融合蛋白诱发豚鼠迟发型超敏反应的实验研究

Study on DTH reaction of guinea-pig induced by recombinant CFP10-ESAT6 antigen of M. tuberculosis

目的探讨以rCFP10-ESAT6融合蛋白为抗原用皮试法评价其对结核分枝杆菌（Mtb）和卡介苗（BCG）致敏豚鼠的鉴别意义。 方法分别用灭活H37Rv和BCG活菌致敏豚鼠．4wk后用不同剂量的rCFP10—ESAT6对同一只豚鼠作皮试，同时设立5IU结核菌素纯蛋白衍生物（PPD）阳性对照及生理盐水（NS）阴性对照，于注射后24、48、72h测量红肿或硬结的直径。 结果以rCFP10-ESAT6为抗原的皮肤试验检测Mtb致敏豚鼠的特异性和敏感性分别为100．0％（21／21）、90．9％（10／11），以PPD为抗原的皮肤试验检测Mtb致敏豚鼠的特异性和敏感性分别为47．6％（10／21）、90．9％（10／11），以rCFP10-ESAT6为抗原的迟发型超敏反应（DTH）中，以1．0μg剂量效果最好．观察时间以皮试后24h效果最好，1．0μg重组融合蛋白在Mtb致敏豚鼠上引发的红肿、硬结比5IU PPD引发的红肿、硬结大。 结论rCFP10-ESAT6能诱导Mtb致敏豚鼠发生DTH，能特异区分Mtb致敏豚鼠及BCG免疫豚鼠，抗原剂量及反应时间是rCFP10—ESAT6诱导豚鼠DTH的主要影响因素。

Aim To evaluate the application of rCFP10-ESAT6 in skin test on guinea pigs sensitized by inactivated H37Rv and BCG. Methods The guinea pigs were sensitized by H37Rv and BCG respectively. 4 weeks later, each guinea pig was intracutaneously injected with different content of rCFP10-ESAT6, NS as negative control and 5IU PPD as positive control. Skin reaction （induration diameter） in guinea pigs was measured at 24 h, 48 h and 72 h after antigen challenge. Results In skin test, the specificity and sensitivity of rCFP10-ESAT6 to detect the Mtb-sensitized guinea pigs was 100.0% （21/21） and 90.9% （10/11） respectively, but that of PPD was 47.6% （10/21） and 90.9% （10/11） respectively. The DTH responses induced by 1.0 /μg rCFP10-ESAT6 were stronger than that of 0.8 μg, 0.6 μg and 0.4 μg. The DTH responses at 24 h were stronger than at 48 h and 72 h. The DTH response to 1.0 μg rCFP10-ESAT6 was stronger than 5 IU PPD. Conclusion The rCFP10- ESAT6 fusion protein can induce DTH responses in Mtb-infected guinea pigs, and it can differentiate Mtb-sensitized and BCG-vaccinated guinea pigs. The dose of antigen and the reaction time were important factors that affect the DTH responses in guinea pigs.