氯霉素全抗原结合比的定量检测研究

 目的 探讨简便、快速定量检测氯霉素（CA）全抗原结合比的方法。 方法 以牛血清白蛋白（BSA）为载体蛋白，采用混合酸酐法将之与氯霉素琥珀酸酯（CA-HS）偶联，合成 CA-BSA全抗原。应用紫外分光光度法对 CA-BSA全抗原进行定性鉴定；分别采用自行改进的三硝基苯磺酸（TNBS）法和紫外分光光度法对 CA-BSA 全抗原结合比进行定量检测。 结果 紫外分光光度法检测显示 CA-BSA 的最大吸收峰高于相同蛋白浓度的 BSA 最大吸收峰，表明 CA-BSA 全抗原偶联成功。TNBS 法测得的 CA-BSA 全抗原结合比为 26，紫外分光光度法测得的 CA-BSA 全抗原结合比为 23，两种方法的检测结果基本一致。 结论 TNBS 法和紫外分光光度法用于氯霉素全抗原结合比的定量检测均具有快速、灵敏、方便和可靠的特点。TNBS 法对半抗原在紫外区无吸收的全抗原结合比测定也可以胜任，因此适用范围更为广泛。

Quantitative determination of combining ratio of complete antigen for chloramphenicol

Objective To find a simple and fast method for quantitative determination of combining ratio of complete antigen for chloramphenicol (CA). Methods The complete antigen for CA-BSA was synthesized by coupling BSA, which was used as the carrier protein, with CA-HS using mixed anhydride method. Then the CA-BSA complete antigen was qualitatively identified by scanning with a UV-VIS instrument. The combining ratio of the complete antigen for CA-BSA was quantitatively determined by using modified 2,4,6-trinitrobenzene 1-sulfonic acid (TNBS) method and UV-VIS methods respectively. Results UV-VIS scanning found that the maximum absorption peak of the CA-BSA was higher than that of the BSA with a same concentration, indicating a successful coupling of a complete antigen for CA-BSA antigen. The TNBS method showed that the combining ratio of the complete antigen for CA-BSA was 26, which was similar to that detected by UV-VIS method (23). Conclusion Both the TNBS and UV-VIS methods are fast, sensitive, convenient, and reliable for quantitative determination of the combining ratio of the complete antigen for CA. The TNBS method can be more widely used, since it is able to determine the combining ratio of the non-UV-absorbing complete antigen.