以新型沙丁胺醇抗原为基础建立ELISA法测定沙丁胺醇残留量(英文)

Preparation of anti-salbutamol antibody based on a new designed immunogen and development of a heterologous indirect ELISA for detection of salbutamol residue

合成了一种新型沙丁胺醇抗原,并以此建立酶联免疫分析法(ELISA)。利用4-氨基苯甲酸作为偶联剂使沙丁胺醇与载体蛋白连接得到免疫抗原,免疫家兔得到沙丁胺醇多克隆抗体。以克伦特罗-卵蛋白代替包被抗原所建立的ELISA法对沙丁胺醇的半数抑制浓度(IC50)为8.97ng·mL?1,该抗体检测克伦特罗时有较强的交叉反应(107%),有望同时测定食物中的克伦特罗和沙丁胺醇残留。用该法测定猪肝中的沙丁胺醇加标量,回收率在70%～99%,批内差<13.3%,批间差<14.3%。利用该法可快速、灵敏地检测沙丁胺醇的药残量,为最终开发商品化的酶联免疫试剂盒打下良好的基础。

To synthesize salbutamol immunogen and develop an enzyme immunoassay （ELISA）,a new salbutamol immunogen was synthesized using 4-aminobenzoic acid as a linker to connect hapten with carrier protein. An enzyme immunoassay based on the antibody prepared was developed and applied to detect salbutamol residue spiked in swine liver. An unusual coating antigen,clenbuterol-ovalbumin （OVA） conjugate instead of salbutamol-OVA conjugate,was used in the immunoassay and the results were discussed based on the structures of related compounds. The antibodies showed high sensitivity in the heterologous assay when using clenbuterol-OVA as a coating antigen,with an IC50 value of 8.97 ng·mL^-1 toward salbutamol. The antibodies prepared showed high cross-reactivity with clenbuterol （107%） and were promising for the simultaneous determination of salbutamol and clenbuterol residues in food and food products. Recovery rates from the salbutamol-spiked swine liver samples were in the range of 70%-99%,while the intra-assay and inter-assay coefficients of variation were 〈13.3% and 〈14.3%,respectively. In summary,the antibodies of salbutamol have been successfully prepared. Sensitive and stable analysis for the detection of salbutamol residues in swine liver was obtained based on the competitive ELISA methods developed in this study.