三种结核分枝杆菌分泌蛋白的抗体制备及其在抗原检测中的应用  

Preparation of three antibodies for secreted proteins of Mycobacterium tuberculosis and their application in detecting of tubercle antigens

作　　者：唐宇龙[1] 杨华[1] 刘湘新[2] 柳斌[2] 秦莲花[1] 郑瑞娟[1] 丁元生[1] 胡忠义[1] 

机构地区：[1]上海市结核重点实验室同济大学附属上海市肺科医院,200433 [2]湖南农业大学动物医学院

出　　处：《中华传染病杂志》2007年第10期597-600,共4页Chinese Journal of Infectious Diseases

基　　金：上海市科委重点专项基金资助课题(06DZ22034)

摘　　要：目的制备结核分枝杆菌CFP10-ESAT-6、MPT84和MPT63-MPT70重组蛋白抗体,探讨其在检测结核抗原中的应用价值。方法应用重组蛋白CFP10-ESAT-6、MPT64和MPT63- MPT70免疫血清,制备包被抗体和标记抗体,建立检测3种蛋白的双抗体夹心法,并将其用于17种分枝杆菌培养上清液和341份临床血清标本的检测。结果CFP10-ESAT-6的最低检出浓度为25 ng/mL,MPT64和MPT63-MPT70均为50 ng/mL。CFP10-ESAT-6检测17种分枝杆菌培养上清液,只有结核分枝杆菌为阳性;而MPT64和MPT63-MPT70检测除结核分枝杆菌阳性外,还各有5种非结核分枝杆菌培养上清液为阳性结果。CFP10-ESAT6、MPT64和MPT63-MPT70检测165例临床诊断的肺结核患者血清标本阳性率分别为33.9%、41.2%和47.9%;检测112例非结核患者血清标本的假阳性率分别为1.8%、7.1%和10.7%;检测64例健康对照的假阳性率分别为0、3.1%和7.8%。结论CFP10-ESAT-6、MPT64和MPT63-MPT70重组蛋白制备的抗体检测血清标本结核抗原的阳性率均不高,但特异性较好。Objective To prepare antibodies of Mycobacterium tuberculosis（MTB） recombinant proteins CFP10-ESAT-6, MPT64 and MPT63-MPT70 and evaluate their application in the detection of tuberculosis antigens. Methods Sera of rabbits, which were immuned by recombinant proteins CFP10-ESAT-6, MPT64 and MPT63-MPT70, were used to prepare enzyme-labelled antibodies and coated antibodies. A double-mAb sandwich enzyme-linked immunosorbent assay（ELISA） method was established to detect the culture supernatant of 17 Mycobacteria strains and 341 clinical serum samples. Results The minimum detectable concentration of CFP10-ESAT-6, MPT64 and MPT63- MPT70 was 25 ng/mL, 50 ng/mL and 50 ng/mL, respectively. When the culture supernatants of 17 Mycobacteria strains were detected by CFP10-ESAT-6, only MTB was positive; and when detected by MPT64 and MPT63-MPT70, MTB as well as five non-tuberculous Mycobacteria was positive. The positive rates of ELISA, based on CFP10-ESAT6, MPT64 and MPT63-MPT70 in detecting the sera of 165 patients with pulmonary tuberculosis were 33.9%, 41.2% and 47.9%, respectively. False positive rate in diagnosis of 112 patients with non-tuberculous were 1.8%, 7. 1% and 10.7G, respectively;were 0, 3.1% and 7.8% in diagnosis of 64 healthy individuals, respectively. Conclusions The positive rates of the method using CFP10-ESAT-6, MPT64 and MPT63-MPT70 recombinant proteins as antibodies to detect tuberculosis antigens in serum are not as high as expected, but the specificities are high.

关 键 词：分枝杆菌 结核 重组蛋白质类 ESAT-6-CFP10融合蛋白 MPT64蛋白 结核分枝杆菌 MPT63-MPT70 

分 类 号：R446.6[医药卫生—诊断学]