双酚A单克隆抗体的制备

摘要

以双酚酸为起始原料,通过3碳间隔臂联接牛血清白蛋白(BSA),制备了双酚A(BPA)人工抗原1：BPA-C3-BSA。以双酚A为起始原料,在其中一个酚羟基位置通过4碳间隔臂联接BSA,制备了BPA人工抗原2：BPA-C4-BSA。分别将这两种抗原免疫Balb/C小鼠,经克隆、亚克隆后获得稳定分泌细胞株BPA1-6E3和BPA2-4C7。对细胞上清液中的抗体进行鉴定,两株细胞所分泌抗体对BPA的IC50值分别为43.76 ng/mL和3.56 ng/mL。进一步鉴定4C7的IgG亚型为IgG1,轻链亚型为κ,抗体亲和常数为6.2×109。抗体与双酚酸的交叉反应率为4.9%,与双酚B的交叉反应率小于0.36%。初步实验表明,该抗体与BPA人工抗原2配对,可用于开发BPA酶联免疫试剂盒。%With diphenolicacid as a starting material,the BPA artificial antigen 1:BPA-C3-BSA was synthesized by coupling BSA to the methylene of BPA via a three-carbon spacer arm. Furthermore,BPA artificial antigen 2:BPA-C4-BSA was synthesized with BPA as a starting material by coupling BSA to one of the phenolic hydroxyl of BPAvia a four-carbon spacer arm. After immuning Balb/Cmice with these two antigens separately,BPA1-6E3 and BPA2-4C7 cell lines that stably secreting BPA antibodies were obtained after clone and subclone. The IC50 of the cell supernatant from two cell lines to BPA were 43. 76 ng/mL and 3. 56 ng/mL respectively. The IgG subtypes of 4C7 is IgG1,light chain subtype is κ. The affinity constant of 4C7 to BPA is 6. 2×109. The cross reactivity with diphenolicacid was 4. 9% and with bisphenol B was less than 0 . 36%. Preliminary experiments show that antibody 4 C7 can react with BPA-C4-BSA,which can be used to develop enzyme-linked immunoassay kit for BPA.