百菌清人工抗原的制备与鉴定

Preparation and Identification of Tetrachloroisophthalonethile Artificial Antigen

以杀菌剂百菌清为起始反应物,通过两步化学反应合成百菌清的衍生物(半抗原);采用活化酯法与混合酸酐法分别将半抗原与载体蛋白(牛血清蛋白、卵清蛋白)进行偶联,制备百菌清的人工抗原,同时通过免疫新西兰大白兔获得抗血清,用酶联免疫吸附法(ELISA)测定其效价。利用红外光谱(IR)、质谱(MS)、核磁共振波谱(NMR)对百菌清半抗原进行表征,并用紫外扫描的方法对人工抗原进行鉴定。结果表明,成功制备出百菌清半抗原及人工抗原,并且半抗原与载体蛋白的偶联比分别为27.86:1和12.32:1,抗血清效价约为1:1.28×104。

In this work, tetrachloroisophthalonethile （TCPN） was derivatized into TCPN-COOH through two-step chemical reactions to prepare a hapten. TCPN artificial antigens, TCPN-COOH-BSA and TCPN-COOH-OVA were prepared from the hapten separately conjugated with carrier proteins, i.e., bovine serum albumin （BSA） and ovalbumin （OVA） by activated ester and mixed anhydride methods. An antisera was obtained through the immunization of New Zealand white rabbits using TCPN- BSA. Antiserum titer was detected by enzyme-linked immunosorbent assay （ELISA）. Meanwhile, TCPN-COOH and its intermediate product TCPN-OH were characterized by IR, MS and NMR methods, and TCPN-COOH-BSA and TCPN- COOH-OVA were qualitatively analyzed by UV/Vis spectrophotometry. TCPN-COOH, TCPN-COOH-BSA and TCPN- COOH-OVA were successfully synthesized, and the coupling ratios between TCPN-COOH and BSA or OVA were 27.86 and 12.32, respectively. The antiserum titer was about 1：1.28 × 104.