米酵菌酸单克隆抗体的制备与鉴定

目的 建立鲜湿米粉中米酵菌酸(bongkrekicacid,BA)的免疫学快速检测方法,制备特异性识别BA的单克隆抗体并进行评价。方法 利用碳二亚胺1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride,EDC]法将BA半抗原与牛血清白蛋白(bovineserumalbumin, BSA)和卵清蛋白(ovalbumin, OVA)载体偶联,分别合成BA免疫原(BA-BSA)和包被原(BA-OVA),用BA-BSA免疫Balb/C小鼠,取免疫效果好的小鼠脾脏与小鼠NS-1骨髓瘤细胞进行融合。采用间接竞争酶联免疫吸附法(indirect competitive enzyme-linked immunosorbent assay, icELISA)进行筛选,筛选出能分泌所需特异性抗体的杂交瘤细胞,利用有限稀释法进行亚克隆得到单株能稳定分泌所需抗体的细胞;采用体内诱生法制备腹水型单克隆抗体。利用正辛酸-饱和硫酸铵法纯化腹水型抗体,通过酶联免疫吸附法测定纯化后的抗体效价。结果 成功合成了BA免疫原BA-BSA和BA包被原BA-OV...

TAN Pan1, WU Xin1,TANG Fei1 , HUANG JianFei2,ZHANG GuiHong2,CHEN Jing2*

objective to establish an immunological method for rapid detection of monoclonal antibody acid in fresh and wet rice noodles, and to prepare and evaluate it. Methods Balb/C mice were immunized with immunogens prepared by Carbodiimide EDC, and the spleen of the mice was fused with NS-1 myeloma cells. The hybridoma cells secreting specific antibody were screened by competitive binding double-positive two-step screening method, and the individual cells were obtained by limited dilution subcloning method, and the ascitic monoclonal antibody was prepared by in vivo induction method. The ascitic type antibody was purified by the octanoic acid-saturated ammonium sulfate method, and the purified antibody was identified by the ELISA method. Results BSA immunogen and OVA coating antigen were successfully synthesized and the phenolphthalein hybridoma cell line babsa 2023 was obtained. The titer of monoclonal antibody was 1 × 105. Conclusion: the method for preparation of rice monoclonal antibody acid with high specificity has been established, which provides a theoretical basis for the development of a rapid immunoassay kit for rice fermentative acid in fresh and wet rice noodles.