结核分支杆菌特异性抗原重组蛋白MPT63 检测系统的建立.docVIP

结核分支杆菌特异性抗原重组蛋白MPT63检测系统的建立 程国平 李克生 张润玲 【摘要】目的建立规范的结核分枝杆菌(mycobacterium tuberculosis,MTB)特异性抗原重组蛋白MPT63酶联免疫吸附试验(enzyme-linked immunosorbent assay, ELISA)检测系统。 方法 采用方阵法确定蛋白包被浓度和酶标二抗浓度，以ROC（Receiver Operating Characteristic）曲线法确定MPT63蛋白 ELISA法检测的cut-off值。探讨规范的ELISA检测系统建立的方法。 结果 ELISA检测系统MPT63蛋白包被浓度1﹕128000，酶浓度1﹕1000时为最佳；在cut-off值为0.35-0.4时检测MPT63蛋白抗体的灵敏度和特异性最为理想，分别是75%和 78.6%。 结论 用方阵法确定蛋白包被浓度和酶标二抗浓度，并以ROC曲线法确立cut-off值是建立规范化的ELISA检测系统的关键环节。 关键词 重组蛋白MPT63；ELISA； ROC曲线； cut-off值 Establishment detection system of recombination protein MPT63 on mycobacterium tuberculosis specific antigen CHENG Guo-ping,Department of Clinical Laboratory, The First Affiliated Hospital of Henan University of Science and Technology,Henan 471000,China LI Ke-sheng Medical Science Research Institute of Gansu province Biotechnology Center，Zhang Run-ling The original Department of Clinical Laboratory ,The Second Affiliated Hospital of Lanzhou University Laboratory 【Abstract】Objective To set up a standard testing system of mycobacterium tuberculosis specific antigen- recombinated MPT63 protein by enzyme-linked immunosorbent assay method. Methods The square matrix method is used to determine the concentration of coated protein and the concentration of enzyme labeling antibody. Receiver Operating Characteristic method is used to determine the cut-off value of ELISA detected MPT63 protein.To approach a establishment of standard ELISA detection system. Results ELISA detected MPT63 protein may attain higher sensibility and specificity when the coated protein is diluted according to 1﹕128000 and the enzyme labeling antibody is diluted by 1﹕1000. Conclusions The critical elements in ELISA detection system are the application of square matrix method to determine the concentration of coated protein and the concentration of enzyme labeling antibody and application the ROC to determine cut-off. Key words Recombination protein MPT63；ELISA;ROC;Cut-off; 结核病（TB）已成为我国乃至世界危害最严重的疾病之一[1]，2000年第四次全国结核病流行病学抽样调查发现,我