氯霉素多克隆抗体的制备及免疫活性研究

Synthesis and identification of artificial antigen for chloramphenicol

为建立氯霉素的免疫分析方法，由氯霉素原药与丁二酸酐反应合成了氯霉素两种半抗原（CAP-HS1、CAP-HS2），分别纯化后经质谱法（MS）分析鉴定结构．纯化的半抗原与牛血清蛋白（BSA）以碳二亚胺法偶联得到人工抗原．将免疫抗原分别免疫免子获得了相应抗体．当包被抗原的浓度为10μg·mL^-1时，抗CAP-HS1和CAP-HS2的冻干粉效价分别为8．39×10^6和6．55×10^4（OD490nm=1．0）．研究了抗原抗体反应的亲和力和竞争抑制能力，其中抗CAP-HS1抗体适合免疫分析，检测线性范围为0．2～100ng·mL^-1，最低检测限IC20为0．239ng·mL^-1，抗CAP-HS1抗体对各类似物如甲砜霉素、氟甲砜霉素的交叉反应率分别为0．025％和0．166％，对其他类抗生素的交叉反应率〈2×10^-5．

In order to establish the immunoassay method of chloramphenicol（CAP）, two haptens （CAP- HS1, CAP-HS2） were synthesized by chloramphenicol and succinic anhydride and identified by ESI-MS. The haptens were conjugated to bovine serum albumin （BSA）, with the modified active ester method and used as immuneogens to produce their corresponding antibodies in rabbits. Titers of antibodies against CAP-HS1 and CAP-HS2 were 8.39×10^6 and 6.55×10^4, respectively, at the concentration of coating antigens 10μg·mL^-1. Based on the affinity and specificity of the antibodies, antibody against CAP-HS1 was adopted in the immunoassay. The lineal range of antibody against CAP-HS1 reacting with chloramphenicol was from 0.2 to 100 ng·mL^-1, the limit of detection （LOD） being 0.239 ng·mL^-1. The percentages of cross-activity to thiamphenicol, florfenicol, and other antibiotics were 0.025%, 0.166%, and less than 2×10^-5%, respectively, in this study.