抗双氯青霉素单克隆抗体的制备与初步鉴定

Preparation and identification of monoclonal antibody against dicloxacillin

采用碳化二亚胺法,将双氯青霉素(dicloxacillin)与牛血清白蛋白偶联制备免疫原,免疫BALB/c小鼠,取免疫小鼠的脾细胞与Sp2/0骨髓瘤细胞融合,间接ELISA筛选,阳性杂交瘤细胞用有限稀释法克隆,获得1株可稳定分泌McAb的杂交瘤细胞5D4-C9-C8;间接ELISA方法测定,其细胞上清抗体效价为1∶300,腹水效价为1∶3.5×105,为IgG1,与其结构类似物邻氯青霉素、苯唑青霉素的交叉反应率分别为17.6%和26.1%,与苄青霉素、羟氨苄青霉素和氨苄青霉素均无交叉反应性。体外传代培养和冻存复苏后抗体分泌稳定。为进一步研制检测双氯青霉素的ELISA试剂盒奠定了基础。

DX-conjugated antigen（BSA-DX） were produced by methods of EDC. The spleen cells of BALB/C mouse immunized by BSA- DX were fused with SP2/0 plasmacytoma cells using PEG4000, and selected cultured with HAT and HT medium. A hybridoma cell line of 5D4-C9-C8 was screened for specificity to DX, and cloned by limited dilution method for 3 times, which secrete high affinity and stable monoclonal antibodies against DX with indirect ELISA. The antibody was characterized by IgGl isotype, the titers of antibody in cell supernatant and ascites were 1：300 and 1：3.5 ×10^5, respectively. The McAb against DX was selective toward Cloxacillin, Oxacillin with cross-reactivity of about 17.6 % and 26.1%, which can be used to detect DX residues by competitive ELISA.