酶联免疫斑点检测斑点形成细胞在结核潜伏感染诊断中的价值

The value of the enzyme-linked immunospot assay detecting spot forming cells in the diagnosis of latent tuberculosis infection

目的探讨酶联免疫斑点检测（ELISPOT）斑点形成细胞（SFC）在结核潜伏感染中的诊断价值。方法研究对象分为：结核组132例，健康对照组44名，有结核暴露史的健康者55名。应用克隆表达的早期分泌抗原靶6KD／培养滤液蛋白10KD融合蛋白（ESAT-6／CFP-10融合蛋白）、早期分泌抗原靶6KD（ESAT-6）进行ELISPOT，检测观察对象的SFC的数量，并对有结核暴露史者进行结核菌素试验（TST）。结果ESAT-6／CFP-10融合蛋白-ELISPOT、ESAT-6-ELISPOT在结核组形成SFC数量（M分别为147．5和27．5）显著高于有结核暴露史健康组形成的SFC数量（M分别为30和10，P值分别为0．000和0．003）；有结核暴露史健康组形成的SFC数量（M分别为30和10）显著高于健康对照形成SFC的数量（M分别为10和5，P均小于0．05）。ESAT-6／CFP-10-ELISPOT与ESAT-6-ELISPOT比较，其与结核暴露史具有更明显的相关性（X^2=6．942，P=0．008），而两种ELISPOT检测均与BCG接种无关；TST与BCG接种显著相关（X^2=8．967，P=0．003）。结论ESAT-6-ELISPOT与结核菌素试验比较，ESAT-6／CFP-10融合蛋白。ELISPOT有可能根据SFC的数量来鉴别结核潜伏感染和活动性结核病，成为诊断结核潜伏感染的更加准确的方法。

Objective To explore the value of Enzyme-Linked Immunospot Assay （ELISPOT） in the diagnosis of latent tuberculosis infection. Methods The peripheral blood mononuclear cells （PBMC） of all participants were co-cultured with the following antigens： Early Secretory Antigenic Target-6/Culture Filtrate Protein-10 fusion protein （ESAT-6/CFP-10 fusion protein ）, Early Secretory Antigenic Target-6 （ESAT-6）, and spot forming cells （SFC） were enumerated by ELISPOT. Tuberculin Skin Test （TST） was also administered to healthy cases that have exposure history to tuberculosis. Results SFC counts after stimulation with ESAT-6/CFP-10 fusion protein and ESAT-6 showed much higher level in active tuberculosis （M = 147. 5 and 27. 5） than that in healthy cases which have exposure history to tuberculosis （M = 30 and lO,respectively）, and SFC counts also much higher in healthy cases which have exposure history to tuberculosis than that in healthy control （ M = 10 and 5, respetively, P 〈 0. 05 ）. ESAT-6/CFP-10 fusion protein-ELISPOT showed much significant correlation than ESAT-6-ELISPOT with M. tuberculosis exposure （X^2 = 6. 942, P = 0. 008 ）, while ESAT-6/CFP-10 fusion protein-ELISPOT and ESAT-6-ELISPOT didn＇t associated with BCG vaccination, TST mainly correlated with BCG vaccination （X^2 =8. 967,P=0.003）. Conclusions ESAT-6/CFP-10 fusion protein-ELISPOT probably offers a more accurate approach than ESAT-6-ELISPOT and TST for identification of individuals who have latent tuberculosis infection. ESAT-6/ CFP-10 fusion protein- ELISPOT may be used to distinguish active tuberculosis and healthy cases with tuberculosis exposure history according SFC count.