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目的制备石杉碱甲单克隆抗体,分析其免疫学特性,建立蛇足石杉中石杉碱甲的ELISA检测方法。方法用石杉碱甲人工抗原免疫Balb/c小鼠,采用杂交瘤技术筛选获4株单克隆抗体3E02、4B01、5D03和4C04,对单克隆抗体纯度、亚型、灵敏度、特异性等进行鉴定,建立ELISA检测方法。结果 4株抗体亚型分别为IgG2b,IgG2b,IgG1和IgG1。ELISA检测方法的标准曲线方程为Y=0.373 6X-0.236,r=0.996 5,线性范围为5~1 000 ng·mL-1,检测限4.387 ng·mL-1。石杉碱甲的平均回收率为99.5%,RSD为0.68%(n=6)。结论 ELISA法快速灵敏,可用于蛇足石杉中石杉碱甲的含量测定。
OBJECTIVE To prepare huperzine A monoclonal antibody and analyze its immunological characteristics to establish an ELISA method for detection of huperzine A in Huperzia serrata. Methods Balb / c mice were immunized with the artificial antigen of Huperzine A, and the monoclonal antibodies 3E02, 4B01, 5D03 and 4C04 were screened by hybridoma technique, and the purity, subtype, sensitivity and specificity of the monoclonal antibodies were identified , Establish ELISA test method. Results The four antibody subtypes were IgG2b, IgG2b, IgG1 and IgG1. The standard curve of ELISA was Y = 0.373 6X-0.236, r = 0.996 5, linear range was 5-1 000 ng · mL-1 and the limit of detection was 4.387 ng · mL-1. The average recovery of huperzine A was 99.5% with a RSD of 0.68% (n = 6). Conclusion The ELISA method is rapid and sensitive and can be used for the determination of huperzine A in Huperzia serrata.