| 摘要: |
| [目的]制备磺胺嘧啶(Sulfadiazine,SD)单克隆抗体(Monoclonal antibody,mAb),并对其免疫学特性进行检测.[方法]用重氮化法将SD偶联于载体蛋白牛血清白蛋白(BSA)和卵清蛋白(OVA)上,合成免疫原BSA-SD和包被原OVA-SD,并用紫外扫描(UV)、SDS-PAGE法进行鉴定.用BSA-SD免疫Balb/c小鼠,间接ELISA和阻断ELISA法筛选细胞融合备用鼠;应用杂交瘤技术建立SD mAb细胞株,用体内诱生腹水法制备SD mAb,对SD mAb的效价、敏感性和特异性等免疫学特性进行鉴定.[结果]BSA-SD人工抗原制备成功,SD 与BSA的分子结合比约为10.3∶1;筛选出6B5-E4、6B5-E9、3A5-B4和2H8-G5 4株敏感特异的杂交瘤细胞,其细胞培养上清液效价分别为1∶810、1∶256、1∶512、1∶512,腹水效价分别为1∶(8.6×106)、1∶(2.5×106)、1∶(5×106)和1∶(5×106).6B5-E4株对SD的半数抑制浓度(IC50)为0.54 μg/L.SD mAb与磺胺一甲基嘧啶交叉反应率为2.7%,与其他磺胺类药物无交叉反应性.[结论]获得了抗SD效价高、敏感、特异的mAb,可用于SD残留的快速检测. |
| 关键词: 磺胺嘧啶 单克隆抗体 免疫学特性 |
| DOI： |
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| 基金项目:国家“十一五”科技支撑计划项目（2006BAK02A21） |
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| Preparation and immunological trait of monoclonal antibody for sulfadiazine |
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| Abstract: |
| 【Objective】The monoclonal antibodis against Sulfadiazine were prepared and their immunological characteristics were identified.【Method】Diazotization was used to conjugate SD to carrier protein BSA and artificial antigen was obtained.The coating antigen OVA-SD was obtained in the same way, ultraviolet（UV）and SDS-PAGE were used to identify SD artificial antigen.BALB/C mice were immunized with BSA-SD,the titre of polyclonal antibody（pAb） was detected by indirect ELISA and blocking ELISA. The hybridoma lines that secrete SD mAb were established with monoclonal antibody hybridoma technology.The immunological characteristics such as titer,sensitivity and specificity of the mAb were studied.【Result】The results showed that SD artificial antigen was synthesized successfully and its conjugation ratio of SD to BSA was about 10.3∶1,three hybridoma cell lines of 6B5-E4、6B5-E9、3A5-B4 and 2H8-G5 were screened for specificity to SD,the indirect ELISA titer of the mAb were 1∶810,1∶256,1∶512,1∶512 and in supernatant,1∶(8.6×106),1∶(2.5×106),1∶(5×106),1∶(5×106) in ascites,the mAb of 6B5-E4 showed good sensitivity with an IC50 of 0.54 μg/L to SD, 2.7% cross-reactivity to Sulfamonomethoxine and little or no cress-reactivity to other compounds. 【Conclusion】 SD mAb of high-titer，sensitivity and specificity has been generated and made it possible to establish immunoassay of SD residues. |
| Key words: sulfadiazine monoclonal antibody immunological characteristic |
