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目的:通过分子佐剂C3d3增强hCGb避孕疫苗的免疫原性。方法:采用分子生物学技术以phCMV1为载体分别构建分泌型、带有6个组氨酸纯化标签的真核表达质粒phCMV1-6His-hCGb-C3d3和phCMV1-6His-hCGb,在CHO细胞中获得稳定、高效表达的重组蛋白,并用镍柱和凝胶过滤层析对其进行分离、纯化。分别用hCGb-C3d3融合蛋白和单用hCGb间隔4周两次免疫生育期雌性BALB/c小鼠,ELISA测定血清中抗hCGb抗体滴度,并对各组小鼠产生的抗血清拮抗hCG诱导的小鼠子宫增重效应进行比较。结果:C3d3使hCGb蛋白疫苗的免疫原性增强1 995倍,hCGb-C3d3融合蛋白免疫小鼠产生的抗血清具有很强的抑制小鼠子宫增重作用。结论:通过分子佐剂C3d3可以大幅提高机体对hCGb的体液免疫应答能力,hCGb-C3d3融合蛋白免疫小鼠产生的抗血清对hCG的生物学作用具有更强的抑制效应。
OBJECTIVE: To enhance the immunogenicity of hCGb contraceptive vaccine by molecular adjuvant C3d3. Methods: The eukaryotic expression plasmids phCMV1-6His-hCGb-C3d3 and phCMV1-6His-hCGb secreting and tagged with 6 histidine were constructed by molecular biology techniques using phCMV1 as vector, respectively, and were stable in CHO cells , Highly expressed recombinant protein, which was separated and purified by nickel column and gel filtration chromatography. Female BALB / c mice were immunized twice with hCGb-C3d3 fusion protein and hCGb alone for 4 weeks respectively. Anti-hCGb antibody titers in serum were measured by ELISA. Anti-hCG-induced antisera Mouse uterine weight gain effects were compared. RESULTS: C3d3 enhanced the immunogenicity of hCGb protein vaccine by 1 995 times. The antiserum raised by hCGb-C3d3 fusion protein had a strong inhibitory effect on uterine weight gain in mice. CONCLUSION: The humoral immune response to hCGb can be greatly enhanced by the molecular adjuvant C3d3. The antiserum raised by hCGb-C3d3 fusion protein has a stronger inhibitory effect on the biological effect of hCG.