利用重组HPV16L1抗原检测宫颈癌抗L1或VLP抗体的对比

为了评价重组大肠杆菌表达的HPV16L1蛋白和重组腺病毒表达的HPV16L1 VLP两种抗原在检测宫颈癌抗 16L1或VLP抗体及在宫颈癌血清学诊断意义上的差别 ,应用PCR技术从宫颈癌组织的DNA中扩增出全长15 35bp的HPV16L1基因片段 ,克隆至 pUC18 T载体中 ,进行DNA测序鉴定。然后 ,将HPV16L1基因克隆至pGEX 2T表达载体中 ,并诱导表达HPV16L1融合蛋白 ,分子量为 83kD ,能被HPV16L1单克隆抗体所识别。经GST柱层析法纯化后 ,与重组腺病毒表达的HPV16L1 VLP分别经酶联免疫吸附 (ELISA)法检测 12份宫颈癌患者和 35份献血员血清。 12例宫颈癌血清标本中 ,抗HPV16L1蛋白的抗体阳性率为 7例 (占 5 8.3% ) ;抗HPV16L1 VLP的抗体阳性率为 8例 (占 6 6 .7% )。经大肠杆菌表达的重组抗原HPV16L1检测为HPV16抗体IgG( )的 7份患者血清 ,利用HPV16L1 VLP试剂盒检测均阳性 ;经大肠杆菌表达的重组抗原检测为HPV16抗体IgG( )的 5份患者血清 ,利用HPV16L1 VLP试剂盒检测有 1份阳性。两者对HPV16抗体的阳性检出率并无显著差异 (P >0 .0 5 )。本实验结果说明HPV16与宫颈癌高度相关 ,利用大肠杆菌表达的重组抗原HPV16L1和HPV16L1 VLP重组抗原检测抗体的敏感性并不受影响。利用重组抗原HPV16L1对宫颈癌的抗体进行定性、定量分析有助于该疾病

Comparison of Detection Antibodies to Human papillomaviruses 16 L1 in the Cervical Cancer People with Different Recombinant Antigen

In order to use major capsid protein L1 of %Human papillomaviruses%(HPV)16 produced in a fused form in %E.coli% and HPV16 L1 VLP produced in recombinant adenovirus in 293 cells as antigen to detecton antibodies of HPV 16 L1 in the cervical cancer people,and compare the serological difference of the two antigen in the diagnosis of cervical cancer,we used PCR to amplify HPV16L1 gene from the cervical cancer, then cloned into pUC18 T. After DNA sequencing, HPV16L1 gene was cloned into pGEX 2T expressing vector, and induced by IPTG to express in %E. coli% as glutathione S transferase L1 (GST L1) fusions and purified to near homogeneity as antigen, together with HPV16 L1 VLP produced in recombinant adenovirus in 293 cells,to test antibodies of human papillomavirus(HPV)16 L1 of 12 cervical cancer and 53 blood donors. The gene derived from the cervical cancer HPV16 genome was 1535 bp in length, and expressed by %E. coli% to the full length 83 kD polypeptide, which recognized by HPV16L1 monocloned antibody. In the 12 cervical cancer sera, there were 7 positive in HPV16L1 antibody (58.3% );while 8 positive in HPV16L1 VLP antibody(66.7%). Among 7 positive in anti HPV16 L1using HPV16L1 protein from the %E.coli% as antigen, all are positive when using HPV16L1 VLP as antigen. While among 5 negative in anti HPV16 L1using HPV16L1 protein from the %E.coli% as antigen, there is 1 positive when using HPV16L1 VLP as antigen. There are no difference between two group as antigens in ELISA detection. (P>0.05) Our research suggested that HPV 16 is highly associated with cervical cancer. The sensitivity of the test is the same whether using HPV16L1 protein from the %E. coli% or HPV16L1 VLP as antigens. To detect HPV16L1 antibody is helpful to diagnosis cervical cancer.