结核分支杆菌MPT63抗原的表达及其血清学诊断价值的研究  

THE EXPRESSION OF MPT63 PROTEIN FROM M. TUBERCULOSIS AND ITS VALUE ON SERODIAGNOSIS OF TUBERCULOSIS

作　　者：吴雪琼[1] 张俊仙[1] 夏湘萱[1] 刘军[1] 李洪敏[1] 

机构地区：[1]解放军309医院结核病研究中心,北京100091

出　　处：《中国现代医学杂志》2001年第4期8-10,共3页China Journal of Modern Medicine

摘　　要：获得结核分支杆菌重组MPT6 3蛋白 ,研究其免疫学特性 ,评价其在结核病血清学诊断中的价值 ,寻找更有效的结核病诊断试剂。方法 :应用基因工程技术表达、纯化MPT6 3蛋白 ,通过Westernblotting和免疫双扩散试验分析其抗原性 ,通过斑点免疫试验、结明试验和结核病一步法检测血清中抗结核抗体。结果 :重组质粒 pET2 4b -MPT6 3测序表明具有正确的编码序列 ,MPT6 3蛋白在大肠杆菌细胞内以可溶性蛋白的形式高效表达 ,表达量占菌体总蛋白的 40 %左右 ,分子量约 16kDa ,Westernblotting和免疫双扩散试验检测抗结核抗体的阳性率分别为 46 .7%和 5 0 % ,结核病一步法检测的阳性率为 70 % ,结明试验的阳性率为 80 %。在34例正常血清中 ,MPT6 3和PPD斑点免疫试验检测抗结核抗体的阳性率分别为 38.2 %和 35 .3 % ,结核病一步法检测的阳性率为 2 3 .5 %。结论 :pET2 4b -MPT6 3大肠杆菌工程菌能以可溶性蛋白的形式高效表达 ,重组的MPT6 3也许可作为结核病血清学诊断的一组抗原之一。Objective:To obtain the recombinant MPT63 protein,to study its immunological characteristics,to evaluate its value in the serodiagnosis of tuberculosis,and to develop more effective diagnostic reagents for tuberculosis.Methods:MPT63 protein was expressed and purified with the genetic engineering techniques.The immunogenicity of MPT63 was analyzed by Western blotting and Double immunodiffusion.64 human sera were detected the antibodies against M.tuberculosis antigens by Dot-immunoblot assay POC-TB assay and MycoDot TM assay.Results:The recombinant plasmid pET24b-MPT63 has a correct gene sequence.The recombinant MPT63 expressed in E.coli exists in the form of soluble protein.SDS-PAGE proved that 40% or so of somatic solubel protein of MPT63 was expressed.Molecular weight was estimated about 16kDa.Western blotting and Double immunodiffusion revealed good antigenicity of MPT63.Of 30 sera from tuberculosis patients,46.7% and 50% were detected antibodies against MPT63 and PPD by Dot-immunoblot assay,respectively.70% were deteced antibodies against M.tuberculosis by POC-TB assay.80% were detected antibodies against LAM by MycoDot TM assay.Conclusions:The recombinant MPT63 protein could be highly expressed in the form of soluble protein in E.coli.It is likely that the recombinant MPT63 can be used as one of multiple antigens in the serodiagnosis of tuberculosis.

关 键 词：抗原 MPT63基因表达 血清学诊断 分支杆菌 结核 

分 类 号：R520.4[医药卫生—内科学] R378.91[医药卫生—临床医学]