丙型肝炎患者血清中丙型肝炎病毒交叉中和抗体的检测

[摘要]目的分析HCV感染者血清中是否存在交叉反应性中和抗体。 方法以分泌表达HCV包膜E2蛋白真核表达质粒转染的293T 细胞培养上清液中的HCV E2蛋白作为检测抗原,建立检测HCV E2抗体的ELISA方法,检测32份HCV抗体阳性的慢性丙肝患者血清,然后用免疫荧光分析血清与HCV全长包膜蛋白表达质粒转染的293T细胞的结合反应,再用5株HCV假病毒(HCVpp)及两株细胞培养产生的HCV(HCVcc)为模型分析血清的病毒中和活性。 结果32份HCV抗体阳性血清中,26份血清可检测出E2抗体,阳性率81.3%。其中HCV RNA阳性的12份血清均为E2抗体阳性,E2抗体水平与HCV RNA水平负相关。HCV E2抗体阳性血清对5株HCVpp以及两株HCVcc的感染性均有不同程度的中和作用,中和活性与E2抗体水平相平行。结论HCV感染可诱导保护性体液免疫应答,丙肝患者血清中存在交叉中和抗体,提示开发能诱导广泛交叉中和抗体的丙肝疫苗具有可行性。

Detection of hepatitis C virus cross-neutralization antibodies in sera of hepatitis C patients

[Abstract]ObjectiveTo investigate the presence of neutralization antibodies in the sera of hepatitis C patients. MethodsA eukaryotic expression plasmid encoding carboxyl terminal-truncated HCV envelope protein 2 (E2) was transfected into human 293T cells. ELISA method was established to examine the anti-E2 antibodies in the sera of 32 hepatitis C patients. The full-length envelope protein expression plasmid was transfected into 293T cells and the reactivity of transfectant with anti-E2 IgG positive sera was analyzed by immunofluorescence. Five strains of HCV pseudotype particle (HCVpp), including H77 (1a genotype), Con-1 (1b genotype), J4 (1b genotype), J6 (2a genotype), and UNK3a (3a genotype), and two strains of cell cultured HCV (HCVcc) were used to assay the neutralization activity of 12 anti-E2 positive serum samples. MethodsELISA results showed that 26 of 32 serum samples were anti-E2 IgG positive, with the positive rate being 81.3%. The 12 serum samples positive for HCV RNA were all anti-E2 IgG, and the virus load was negatively correlated with the anti-E2 antibody level. The anti-E2 positive sera could neutralize the infections of five strains of HCVpp and two strains of HCVcc to various extents, and the neutralization activity was consistent with the anti-E2 antibody level. MethodsIt is suggested that HCV infection can induce protective humoral immune response, and cross-neutralization antibodies against HCV are present in sera of hepatitis C patients, which indicates a feasibility for developing vaccines that can induce broadly-reactive neutralizing antibodies.