人参皂苷Rh₁人工抗原的合成及免疫原性分析

目的：合成人参皂苷Rh₁（GsRh₁）的人工抗原，为获得其单克隆抗体奠定基础。方法：采用高碘酸钠氧化法分别合成GsRh₁免疫抗原（GsRh₁-BSA）和包被抗原（GsRh₁-OVA）；用紫外光谱法和薄层色谱法鉴定人工抗原的合成是否成功；利用GsRh₁-BSA免疫小鼠制备血清多抗；通过优化封闭液和包被原工作浓度，建立酶联免疫吸附分析法（ELISA）的最佳工作条件，并检测血清中抗体效价及特异性。结果：紫外光谱和薄层色谱法鉴定结果表明，GsRh₁与BSA/OVA偶联成功，免疫后小鼠血清中产生了能特异性地与GsRh₁结合的抗体，效价达到1：32000，线性范围为10~10000 ng·mL^-1，经分析确定ELISA的最佳包被浓度为1：4000，最佳封闭液为10 mg·mL^-1的明胶溶液。结论：成功合成了GsRh₁人工抗原，并诱发小鼠产生了抗GsRh₁的抗体，可用于下一步单克隆抗体的制备。

Objective: To synthesize the artificial antigen of ginsenoside Rh₁(GsRh₁) for preparing monoclonal antibody (MAb). Methods: The immunizing antigen (GsRh₁-BSA) and coating antigen (GsRh₁-OVA) were synthesized by sodium periodate oxidation method.The synthesis was characterized by ultraviolet (UV) spectrometry and thin layer chromatography (TLC).The appropriate concentration of GsRh1-OVA and the optimal blocking buffer were chosen to establish the best working conditions for the enzyme-linked immunosorbent assay (ELISA) method.The titer and specificity of the antibody in serum of immunized mice were detected respectively by indirect enzyme-linked immunosorbent assay (iELISA) and indirect competitive enzyme-linked immunosorbent assay (icELISA). Results: According to the UV spectrometry and TLC,the GsRh₁ was successfully conjugated with BSA and OVA.The antibody obtained from immunized mice could bind to GsRh₁ specially and the titer was up to 1:32000.The linear range was between 10 ng·mL^-1and 10000 ng·mL^-1. The analysis indicated that the appropriate dilution ratio of GsRh₁-OVA was 1:4000 for ELISA.The optimal blocking buffer was 10 mg·mL^-1gelatin. Conclusion: The artificial antigen of GsRh₁ was successfully synthesized,and the ant-GsRh₁ antibodies which can be used for further preparation of MAb were generated.