氨苄青霉素人工抗原的合成及单克隆抗体的制备

Synthesis of ampicillin artificial antigen and preparation of anti-ampicillin monoclonal antibodies

用酸酐法将氨苄青霉素(Ampicillin,AMP)偶联于载体蛋白牛血清白蛋白(BSA)和匙孔血蓝蛋白(Keyhole Limpet Hemocyanin,KLH),合成了免疫抗原AMP-KLH和检测抗原AMP-BSA,并进行了鉴定;用免疫抗原AMP-KLH免疫BALB/c小鼠,应用杂交瘤技术筛选出了3株分泌抗AMP单克隆抗体的杂交瘤细胞株。特性分析显示,单克隆抗体2C3、4E5和4C7的腹水效价分别为1∶1.02×106、1∶5.12×105和1∶5.12×105,其中2C3的AMP半数抑制浓度(IC50)为9.3 ng/m L,与青霉素类抗生素有明显交叉反应,其他类别抗生素的交叉反应率均小于0.01%。

Ampicillin （Amp） was conjugated to carrier-protein bovine serum albumin （BSA） or Keyhole Limpet Hemocyanin （KLH） to synthesize the immnunogen KLH-Amp and the detection antigen BSA-Amp by acid anhydride method.KLH-Amp was used to immunize BALB/c mice,and three hybridoma cell lines secreting monoclonal antibodies （mAbs） against Amp were developed using hybridoma technology.The titers of the ascitic fluid （2C3,4E5 and 4C7） were determined as 1 ∶ 1.02×10^6,1 ∶ 5.12×10^5 and 1 ∶ 5.12×10^5 in ELISA,and for 2C3,the half maximal inhibitory concentration （IC50） of Amp were 9.3 ng/mL in the inhibitive ELISA.The Amp mAbs showed the significant cross-reactivity to the penicillin antibiotics like carbenicillin,penicillin G and amoxicillin,whereas no obvious cross-reactivity to other antibiotic was found.