摘要
目的分析丙型肝炎病毒（ＨＣＶ）ＮＳ５蛋白的抗原性，研究抗－ＮＳ５抗体的性质，动态变化规律及临床诊断意义。方法用Ｇｏｌｄｋｅｙ程序对ＮＳ５蛋白的抗原决定簇进行预测分析，以ＲＴ－ＮｅｓｔｅｄＰＣＲ方法扩增ＮＳ５基因片段，并进行核酸序列分析。通过基因体外重组技术克隆表达目的基因。用所获重组蛋白建立ＥＩＡ，对３０例ＰＴ－ＨＣ患者的系列血清进行抗体、丙氨酸转氨酶（ＡＬＴ）和ＨＣＶＲＮＡ检测和分析。结果所获ＮＳ５区基因与ＨＣＶⅡ／１ｂ相应区段核苷酸及氨基酸的同源性分别为９１．８％和９２．４％。将该片段插入ｐＧＥＸ表达载体中，转化ＢＬ２１（ＤＥ３），通过诱导表达，在分子量５００００左右处出现一条对照菌没有的表达蛋白带，表达的融合蛋白占菌体总蛋白的２１．４％。Ｗｅｓｔ－ｅｒｎＢｌｏｔ结果表明，重组蛋白可与丙型肝炎患者血清发生特异性反应。系列血清的检测表明：抗－ＮＳ５抗体一般出现较晚，阳转时间为１８３±１６９天，抗体持续时间相对较短，抗－ＮＳ５抗体的出现与ＡＬＴ的升高有一定的关系。ＮＳ５区抗体的动态变化可分为：持续性抗－ＮＳ５阳性、间歇性抗－ＮＳ５阳性、一过性抗－ＮＳ５阳性和２年内抗－ＮＳ５持续阴性四种类型。结论抗－ＮＳ５出?
Objective To study antigenicity of HCV NS5 protein and dynamic changes of anti NS5 in post transfusion hepatitis C (PT HC). Methods Epitopes of HCV NS5 protein were analyzed by Goldkey Program. NS5 gene fragment was amplified by reverse transcription and polymerize chain reaction (RT PCR) from sera of PT HC patients. Sequence analysis was performed, and recombinant strain was constructed. Series sera from PT HC were detected for anti NS5, ALT and HCV RNA. Results The homology of nucleotide and amino acid with genotype Ⅱ in the same region was 91.8% and 92.4%, respectively. SDS PAGE analysis showed an expressing band around 50kD, the fusion protein represented about 21.4% of total bacterial protein. Western blot result proved the expressing band could specifically react with sera from hepatitis C patients. Detection results of series blood samples from PT HC showed that the antibody against NS5 appeared relatively late, the positive conversion time was 182.9±168.5 day. Dynamic changes of anti NS5 were correlated with serum ALT in most cases. The types of dynamic change of anti NS5 were passing positive; intermittent positive; persistent positive, and persistent negative in two years.Conclusion Antibody against NS5 may reflect the disease activity to some extent. It appears relatively late, and is of no value in early stage diagnosis
出处
《中华医学杂志》
CAS
CSCD
北大核心
1998年第3期183-186,共4页
National Medical Journal of China
基金
国家863高科技计划
