摘要
用与牛血清蛋白(BSA)交联的克百威人工抗原(BFNB-BSA)免疫的BALB/C鼠脾细胞与SP2/0鼠骨髓瘤细胞融合,经筛选、克隆,得到1株能稳定分泌克百威抗体的单克隆细胞株(1E8)。鉴定结果表明,1E8的抗体类型及亚类均为IgG1,其轻链为κ链。制备单克隆抗体腹水,腹水的间接ELISA(酶联免疫吸附分析)效价在1×10-6以上。直接ELISA线性回归方程为y=15.70lg(x)-5.879 5,r2=0.990 4,抑制中浓度IC50=35.1μg.L-1,最低检测限IC20=5.2μg.L-1。该单克隆抗体与克百威特异性结合反应的50%抑制质量浓度为35.1μg.L-1。除丁硫克百威以外与其他克百威结构类似物无交叉反应。
High affinity monoclonal antibodies （MAb） against carbofuran was produced by using 500 mg·mL^-1 PEG4000 to do cells fusion, and using HAT （H, hypoxanthine; A, aminopterin; T, thymidine） culture medium for screening and limiting dilution for cell sub-cloning. Hybridoma lines were screened for specificity to carbofuran by an Enzyme-linked Immunosorbent Assay （ELISA）. One hybridoma cell line （1E8） secreting monoclonal antibody （MAb） against carbofuran was produced by fusing mouse myeloma cells （SP2/0） with spleen cells from BALB/C which immunized by the artificial antigen conjugated with bovine serum albumin （BSA）. The antibodies of hybridoma culture supernatant had no reaction with BSA and Ovalbumin （OVA）. Isotype and subclass of the monoclonal cell line （1E8） showed that it belonged to IgGl. The light chain of the MAb was identified to be K. The MAb obtained could specifically react with carbofuran with almost no cross-reactivity to carbofuran analogues except carbosulfan, with the titres of ascitic fluids up to 1 ×10^-6 by indirect ELISA. Ascites antibodies generated by hybridoma of 1E8 cells were purified. Inhibition rate studies showed that the detection limit of the ELISA was 5.2 μg· L^-1, with the regression equation of direct ELISA y=15.701g （x）-5.879 5, r^2=0.990 4, IC50 value being 35.1 μg·L^-1. The MAb can be used to prepare the reagents for analyzing carbofuran residue.
出处
《农业环境科学学报》
CAS
CSCD
北大核心
2006年第5期1276-1280,共5页
Journal of Agro-Environment Science
基金
国家自然科学基金(30370944)
