细菌受体分析在禽类血清四环素族残留物检测中的应用

Application and Evaluation of Bacterial Receptor Assay for Multi-tetracycline Residues in Live Poultry Serum

目的:以商售 Charm 细菌四环素族受体与四环素族的特异性结合反应为基础,应用其对活体禽类动物血清中四环素族抗生素残留总量进行快速筛选,重点对方法的灵敏度、选择性、基质影响和准确性等进行综合评价。方法:血清经缓冲液稀释,其中的四环素族抗生素残留与~3H 标记的金霉素竞争结合特异性细菌受体。竞争反应温育时间5 min,温度35℃。3300r·min^(-1),5 min 离心分离倾去未结合的游离化合物,复合物沉淀用水溶解,加入闪烁液混匀后放置1 min,进行[~3H]通道的60s 液体闪烁计数。结果:在大于95%的置信水平,血清中四环素、土霉素和金霉素的检测低限分别为80,55,15μg·L^(-1);样本测试表明筛选水平为200μg·L^(-1)时,假阴性率为0,阳性检出结果中假阳性率不超过15%。结论:与色谱法相比,其快速、灵敏、选择性高,可对活体禽类动物血清中四环素族残留物进行高通量筛选。

Objective：A commercially available Charm bacterial receptor based assay kit was applied for the fast screening of total multi -tetracyclines （TCs） residues in live poultry animal serum, especially focused on the comprehensive evaluation of the sensitivity, selectivity, matrix effect and accuracy of the receptor based screening. Method：The ^3H labeled chlortetracycline tracer was added to animal serum which has been properly diluted by EDTA - Mcvallain buffer,it took 5 min for the tracer drug and serum TCs residues to compete for limited binding sites of bacterial receptor at 35℃. After binding reaction and centrifugation,the TCs -receptor complex deposit was redissolved and scintillation liquid was added, after votex mixing and waiting 1min, the scintillation counter was measured at [^3H] channel. Results：The detection limit for tetracycline, oxytetracycline and chlortetracycline residues in serum were 80,55 and 15μg · L^-1 respectively with a minimum 95% confidence. At the screening level 200μg · L^-1,test data showed the false negative result was zero and the false positive rate in all positive results was under 15%. Conclusion： Compared with chromatography method the receptor assay was rapid ,highly sensitive and selective enough to be used for the high - throughput screening of TCs residues in live poultry animal serum.