磺胺嘧啶单克隆抗体的制备与鉴定

Preparation and Identification of Anti-sulfadiazine Monoclonal Antibody

采用重氮法同时制备结合比分别为9.0、8.0、3.7的3种磺胺嘧啶免疫抗原,并将其免疫BALB/c小鼠,其中结合比为8.0的一组免疫效果最佳。获得一株稳定分泌抗磺胺嘧啶单克隆抗体的杂交瘤细胞(1F6),体内诱生法制备腹水。间接ELISA测定腹水效价为1:1.28×106,建立间接竞争ELISA标准曲线,其检测下限为10.4ng/mL,线性范围为14.8～421.5ng/mL,IC50值为79.1ng/mL,与磺胺二甲嘧啶、磺胺间甲氧嘧啶、磺胺对甲氧嘧啶交叉反应率分别为0.3%、0.99%、4.5%,与磺胺甲噁唑、磺胺喹恶啉、对氨基苯甲酸、磺胺、对氨基苯磺酸未见有交叉反应。经鉴定,单克隆抗体免疫球蛋白为鼠IgG的2b亚类。

In order to generate a monoclonal antibody（McAb） against sulfadianzine（SD）,three separate immunogens with SD/BSA conjugation ratios of 9.0,8.0 and 3.7 were prepared by diazotization method and BALB/c mice were then immunized with the immunogens.An optimum immunization was observed in BALB/c mice immunized with the immunogen with middle SD/BSA conjugation ratio.A McAb（1F6） with high affinity to SD was obtained by hybridoma technique.The titer of the McAb was 1：1.28 ×106 in ascites according to the indirect ELISA determination.IC50 and the lowest detectable limit（LDL） of the McAb in linear range of 14.8-421.5 ng/mL for SD were 79.1 ng/mL and 10.4 ng/mL,respectively.The cross-reactivity ratios of the McAb against sulfamethazine,sulfamonomethoxine and sulfamethoxydiazine were 0.3%,0.99% and 4.5%,respectively,while no cross-reactivity to sulfamethoxazole,sulfaquinoxaline,4-aminobenzoic acid,sulfanilamide or sulfanilic acid was observed.The sub-class of the McAb was identified as IgG2b.