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目的:对华支睾吸虫跨膜蛋白63c基因进行克隆、表达,并通过感染者抗体亚类分析评价其免疫学诊断效果。方法:根据Cs TMP 63c(Gen Bank序列号:AF093242)基因序列,拼接其多个跨膜区片段,人工合成新基因Cs TMP 63o,制备抗原免疫小鼠,检测抗体效价,建立抗体检测系统,初步分析感染者针对Cs TMP 63o的特异性抗体亚类。结果 :从Cs c DNA文库中筛选出Cs TMP 63c基因,全长2 013 bp,由大肠杆菌表达、纯化,得到分子量为38 k D的重组蛋白。免疫大鼠后,抗体效价较低;感染者的抗体亚类分布以IgG4类为主,与各项常规生化和血液指标未发现有显著相关性。结论 :Cs TMP 63o在大肠杆菌中高效表达,具有较好的免疫原性,但血清诊断价值不理想。
OBJECTIVE: To clone and express the transmembrane protein 63c gene of Clonorchis sinensis, and to evaluate its immunological diagnosis by analyzing the antibody subtype of infected individuals. Methods: According to the gene sequence of Cs TMP 63c (GenBank accession No: AF093242), multiple transmembrane segments were spliced and a new gene, Cs TMP 63o, was synthesized. The antigens were immunized to detect the antibody titers and the antibody detection system was established. Preliminary analysis of infected patients against Cs TMP 63o specific antibody subclasses. Results: The Cs TMP 63c gene was screened from Cs c DNA library. The full length cDNA was 2 013 bp and was expressed in Escherichia coli and purified. The recombinant protein with molecular weight of 38 kD was obtained. After immunization, the antibody titer was low. The distribution of antibody subtype was mainly IgG4. There was no significant correlation with the conventional biochemical and blood parameters. Conclusion: Cs TMP 63o is highly expressed in Escherichia coli and has good immunogenicity, but the serum diagnostic value is not satisfactory.