摘要
为了在大肠杆菌中表达具有良好免疫反应性的HIV-1gp41重组抗原,本实验运用基因工程技术,经PCR扩增gp41的主要抗原表位序列,BamHⅠ、XhoⅠ双酶切后与E3质粒连接,转化克隆宿主菌DH5α,再提取重组质粒进一步转化表达宿主菌BL21(DE3),经IPTG诱导表达重组蛋白,纯化后标记HRP,通过双抗原夹心酶联免疫方法检测其免疫反应性和特异性。结果表明,获得的HIV-1gp41重组抗原能够与相应抗体特异性结合,与多种无关抗体间无交叉反应,对825份HIV阴性标本检测无错检。检测结果说明该重组抗原具有良好的免疫反应性,在HIV-1抗体诊断试剂中具有潜在的应用价值,为进一步研究gp41抗原奠定了基础。
To express HIV-1 transmembrane glycoprotein gp41 that has fine immunoreactivity in prokaryotic system of E.coli,the target DNA sequence encoding for major epitopes of HIV-1 gp41 was amplified through PCR.The product was digested by BamHⅠand XhoⅠ,and inserted into E3 vector.Then the joint product was transported into cloning host DH5α from which we extracted recombinant plasmid that was further transported into BL21（DE3） host.The recombinant protein was expressed when induced with IPTG.And after being purified and labelled with HRP,the immunoreactivity and specificity of the protein was evaluated through double-antigen sandwich ELISA.Results proved that,the HIV-1 gp41 recombinant antigen bound corresponding antibody specifically,and had no cross reactivity with various irrelevant antibodies and showed no false positive results when testing with 825 HIV negative specimens.So,this recombinant antigen has fine immunoreactivity and potentcial application value in HIV antibody detection kit.
出处
《现代免疫学》
CAS
CSCD
北大核心
2012年第3期230-233,共4页
Current Immunology
基金
上海市科学技术委员会科研计划项目(09DZ2251200)
