摘要：
Objective To compare the antibody expression of three recombination Mycobacterium tuberculosis (MTB) protein antigens (MTB relative molecular mass 38 000 short as 38 kD, MPT64 and HBHA)in the peripheral blood serum, and comprehensively assess their performances in serologic diagnosis of tuberculosis (TB).Methods Seventy eight active tuberculosis (ATB) cases and 36 latent tuberculosis infection (LTBI) cases were selected from Beijing Chest Hospital as case groups, during July 2012 to October 2013.And 31 healthy control (HC) cases were recruited from the TB physical examination in Beijing Chest Hospital at the same period.We used these purified protein generated by our laboratory as antigens to measure the expression level of antibodies in serum by ELSIA, then compared the difference between ATB, LTBI and HC by statistical method.Sensitivity, specificity and diagnostic efficiency were calculated after getting the receiver operating characteristic (ROC) curve analysis with the results of Enzyme-linked immunosorbent assay (ELISA).Furthermore, comprehensive diagnosis performance and application values of these proteins were evaluated.Results The absorbance (A)450 nm value of 38 kD measured by ELISA was higher in ATB group (0.343±0.120) than those in LTBI group (0.221±0.102) and HC group (0.143±0.097).The difference showed statistically significance (t=3.07, P<0.05).The A value of MPT64 measured by ELISA was higher in ATB group (0.234±0.102) than those in LTBI group (0.198±0.087) and HC group (0.123±0.075).The difference showed statistically significance (t=3.79, P<0.05).The A value of HBHA measured by ELISA was higher in ATB group (0.263±0.113) than those in LTBI group (0.188±0.091) and HC group (0.148±0.078).The difference showed statistically significance (t=2.70, P<0.05).Areas under the curve (AUC) of 38 KD, MPT64 and HBHA antibodies which used to distinguish ATB and LTBI were 0.78, 0.61 and 0.62;the sensitivity were 71.79%, 62.82% and 52.56% separately;and the specificity were 72.22%, 58.33% and 69.44% separately.AUC which used to distinguish ATB and HC were 0.91, 0.81 and 0.79;the sensitivity were 87.17%, 69.23% and 73.08% separately;and the specificity were 80.65%, 74.19% and 74.19% separately.AUC which used for distinguish LTBI and HC were 0.63, 0.75 and 0.69;the sensitivity were 58.33%, 77.78% and 63.89% separately;the specificity were 64.52%, 51.61% and 74.19% separately.Conclusion The three proteins possessed good immune-reactivity characteristics, and the expression level of their antibodies showed significantly difference in different patient groups.The assessment of diagnostic efficiency for TB indicated that through detecting antibodies expression levels in peripheral blood of TB patient based on ELISA is an assistant method for diagnosis of TB in clinical laboratory.%目的 对结核分枝杆菌相对分子质量为38 000(以下采用38 kD表示)、线粒体通透性转换蛋白64 (mitochondrial permeability transition 64,MPT-64)和肝素结合血凝素(heparin-binding haemagglutinin,HBHA)蛋白在外周血中抗体的表达水平进行检测和比较,并对它们在结核病血清学方面的诊断价值进行评估.方法 选取2012年7月至2013年10月北京胸科医院收治的肺结核患者作为活动性肺结核(ATB)组,共78例;选取同期36例潜伏结核感染(LTBI)患者作为LTBI组;同期在北京胸科医院进行体检的31名健康人群(HC)作为HC组.将本实验室前期纯化的38 kD、MPT64和HBHA作为抗原应用于免疫学的检测,采用酶联免疫吸附实验(ELISA)检测血清中三种蛋白抗体的表达水平,经过统计学处理后,分别比较ATB、LTBI和HC三组之间的差异,并结合受试者工作特征曲线计算各蛋白抗原的敏感度和特异度,评价三种蛋白血清抗体的临床诊断价值.结果 ELISA检测ATB组38 kD蛋白抗体的吸光度(A)450 nm处的A值(0.343±0.120)高于LTBI组(0.221±0.102)和HC组(0.143±0.097),差异有统计学意义(t=3.07, P<0.05);MPT64抗体的A450值(0.234±0.102)高于LTBI组(0.198±0.087)和HC组(0.123±0.075),差异有统计学意义(t=3.79, P<0.05);HBHA抗体的A450值(0.263±0.113)高于LTBI组(0.188±0.091)和HC组(0.148±0.078),差异有统计学意义(t=2.70, P<0.05).以38 kD、MPT64和HBHA蛋白抗体表达水平从LTBI组鉴别ATB组时的受试者工作特征(ROC)曲线下面积(AUC)分别为0.78、0.61和0.62,敏感度分别为71.79%、62.82%和52.56%,特异度为72.22%、58.33%和69.44%;从HC组鉴别ATB组时的AUC分别为0.91、0.81和0.79,敏感度分别为87.17%、69.23%和73.08%,特异度为80.65%、74.19%和74.19%;从HC组鉴别LTBI组AUC分别为0.63、0.75和0.69,敏感度分别为58.33%、77.78%和63.89%,特异度为64.52%、51.61%和74.19%.结论 MTB 38 kD、MPT64和HBHA蛋白都有较好的免疫反应性,3种蛋白抗体在不同患者人群血清中的表达水平有差异,对肺结核诊断价值的评估结果表明采用ELISA对外周血中MTB蛋白抗体进行检测可作为结核病临床检测诊断的辅助方法.
