人巨细胞病毒糖蛋白B抗原表位在大肠杆菌中的表达

Expression of human cytomegalovirus glycoprotein B epitope

目的:利用大肠杆菌表达人巨细胞病毒(HCMV)糖蛋白B抗原表位AD1和AD2,对表达产物进行纯化,通过ELISA方法检测表达产物与感染HCMV的人血清之间的特异结合反应,探讨其用于临床检测的可行性。方法:以HCMV病毒基因组为模板,PCR扩增AD1和AD2基因,构建重组表达载体pET32-NusA-AD1和pET32-NusA-AD2,在大肠杆菌BL21(DE3)pLysS中利用IPTG诱导表达,通过Ni柱亲和纯化获得NusA-AD1和NusA-AD2,ELISA方法检测NusA-AD1和NusA-AD2与感染HCMV的人血清之间的特异性结合反应。结果:可溶性表达并纯化得到融合抗原NusA-AD1和NusA-AD2。在8份已确定为HCMV IgG阳性人血清标本中,融合抗原NusA-AD1能够与5份血清发生反应,融合抗原NusA-AD2能够与6份血清反应。结论:NusA-AD1和NusA-AD2能够部分检测HCMV感染的人血清。

Objective：To clone and express the epitope AD1 and AD2 of HCMV glycoprotein B（gB） in Escherichia coli, to purify the expression product and to detect sera from HCMV positive patients by ELISA assay for further clinical study, nethods：AD1 and AD2 genes were cloned by PCR using HCMV genome DNA as template and the recombinant expression vectors pET32-NusA-AD1 and pET32-NusA-AD2 were constructed. E. coli BL21 （ DE3 ） pLysS bearing the plasmids was induced with IPTG for protein production. The proteins NusA-AD1 and NusA-AD2 were purified by Ni^2＋ affinity chromatography and used to detect HCMV positive human sera by ELISA method. Results： The recombinant antigens Nu- sA-AD1 and NusA-AD2 were expressed solubly. Of eight HCMV-IgG positive serum samples, five were found positive by NusA-AD1, and six were positive by NusA-AD2. Conclusion： Most HCMV positive human sera could be detected by recombinant antigens NusA-AD1 and NusA-AD2.