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目的研究人类乳头瘤病毒18型E7全长基因(HPV18 E7)在原核表达系统中的表达,及其作为特异性抗原在宫颈癌血清抗体检测中的意义,为HPV感染与肿瘤相关的血清学诊断研究奠定基础。方法PCR法扩增HPV18 E7全长基因,构建重组质粒pET32a(+)/HPV18 E7,转化表达HPV18 E7融合蛋白后经纯化、SDS-PAGE和Western blot分析鉴定。再以该融合蛋白作为诊断抗原,间接ELISA法检测宫颈癌、尖锐湿疣及健康对照三组血清特异性IgG抗体水平。结果HPV18 E7融合蛋白表达量约占菌体总蛋白的35.00%;宫颈癌、尖锐湿疣及健康对照组的血清特异性抗体均值分别为0.604±0.328,0.287±0.202和0.342±0.225,三组间差异有显著性意义(F=32.771,P<0.01);宫颈癌组与尖锐湿疣组及健康对照组的HPV18E7均值比较,差异均有显著性意义(P均<0.01),而尖锐湿疣组与健康对照组抗体均值比较,差异无统计学意义(P>0.05)。结论HPV18 E7融合蛋白具较强的抗原性,对宫颈癌诊断试剂的研究具有应用价值。
Objective To study the expression of human papillomavirus 18 (E18) E7 gene in prokaryotic expression system and its significance as a specific antigen in the detection of serum anti-human cervical cancer antibody. Research laid the foundation. Methods The full length HPV18 E7 gene was amplified by PCR, and the recombinant plasmid pET32a (+) / HPV18 E7 was constructed. The HPV18 E7 fusion protein was transformed and expressed, then identified by SDS-PAGE and Western blot. The fusion protein was used as a diagnostic antigen, and the serum levels of specific IgG antibodies in cervical cancer, genital warts and healthy controls were detected by indirect ELISA. Results The expression of HPV18 E7 fusion protein accounted for about 35.00% of the total bacterial proteins. The mean serum specific antibodies of cervical cancer, condyloma acuminatum and healthy controls were 0.604 ± 0.328, 0.287 ± 0.202 and 0.342 ± 0.225, respectively (F = 32.771, P <0.01). There was significant difference in the average of HPV18E7 between cervical cancer group and condyloma acuminatum group and healthy control group (all P <0.01), but there was no difference between genital warts group and healthy control group There was no significant difference between the mean of antibody group (P> 0.05). Conclusion The HPV18 E7 fusion protein has strong antigenicity and is valuable for the diagnosis of cervical cancer diagnostic reagents.