重金属铅多克隆抗体的制备及鉴定  

Preparation and Identification of Anti-Lead-chelate Complex Polyclonal Antibody

作　　者：翟一凡[1] 向军俭[1] 唐勇[1] 郭常伟[1] 刘小锋 杨红宇[1] 

机构地区：[1]暨南大学抗体工程中心,广东广州510632 [2]广州市千江企业有限公司,广东广州510632

出　　处：《食品科学》2010年第13期180-183,共4页Food Science

基　　金：国家"863"计划项目(2008AA10Z427);广州市天河区科技计划项目(085G079)

摘　　要：目的:合成铅离子完全抗原,并对其进行鉴定;用合成的完全抗原免疫Balb/c小鼠制备多克隆抗体,鉴定效价和特异性。方法:用双功能螯合剂p-SCN-Bn-DTPA将Pb2+与载体蛋白BSA和OVA偶联在一起,合成免疫抗原(Pb-DTPA-BSA)和检测抗原(Pb-DTPA-OVA),并通过SDS-PAGE、紫外扫描、电感耦合等离子体(ICP)检测抗原中Pb2+含量,以及通过免疫Balb/c小鼠,采集血清进行间接ELISA和竞争ELISA检测。结果:两种合成的抗原相对于各自的载体蛋白均出现了蛋白条带滞后,紫外吸收峰偏移等现象;免疫的4号和5号Balb/c小鼠的抗血清效价达1:400000;与OVA无交叉反应,ELISA检测IC50值为1.5ng/mL,与Fe3+有近5%的交叉反应,与其他重金属离子的交叉反应均小于1%。结论:铅离子多克隆抗体制备成功。Objective： BAS and OVA were individually conjugated with lead-chelate complex （Pb-DTPA） to prepare 2 antigens, namely Pb-DTPA-BSA, immunogen and Pb-DTPA-OVA, detective antigen. The synthesized antigens were then used to immunize BALB/c mice for preparing polyclonal antibodies. Finally, the antisera withdrawn from immunized BALB/c mice were assessed for their titers and specificities. Methods： The artificial antigens were identified by SDS-PAGE and scanned with ultraviolet spectrophotometer. The Pb2＋ concentrations of the synthesized antigens were determined using inductively coupled plasma （ICP） spectrophotometer. Indirect ELISA and competitive ELISA were used to determine antiserum titer and specificity, respectively. Results： Compared with carrier proteins, the synthesized antigens exhibited lagged protein bands in SDS-PAGE pattern and blue shift of UV absorption peaks. The antiserum titers of the fourth and fifth mice reached up to 1：400,000 without cross-reactivity with OVA. The IC50 determined by IC-ELISA was 1.5 ng/mL and the cross reactivity with the other heavy ions were less than 1% although the cross-reactivity with Fe3＋ was close to 5%. Conclusion： The above results suggest that anti-lead- chelate complex polyclonal antibodies are prepared successfully.

关 键 词：铅离子 合成抗原 ELISA 多克隆抗体 

分 类 号：Q819[生物学—生物工程]