基于免疫磁珠的胶体金免疫层析法快速检测水产品中地西泮残留

本研究针对水产品中地西泮残留的问题,建立了一种快速、灵敏的免疫磁珠-胶体金免疫层析法。采用链霉亲和素磁珠与生物素化地西泮单克隆抗体偶联制备的免疫磁珠,快速富集、分离水产品中地西泮残留,并结合胶体金免疫层析法进行半定量检测。结果表明,经过优化后试剂卡最佳工艺为金标抗体标记量25 μg/mL、T线抗原包被量0.6 mg/mL、C线羊抗鼠二抗包被量1.5 mg/mL、金标喷量3.0 μL/cm,地西泮加标浓度为1.0、5.0、10.0和15.0 μg/kg时回收率为78.34%~90.40%,相对标准偏差(RSD)为5.03%~8.96%。本方法特异性强、稳定性好,可在25 min内完成单个样本检测,对水产品中地西泮残留检出限为0.5 μg/kg,优于常规前处理法。经验证,对40份水产品样本检测结果与GC-MS法结果一致,证明免疫磁珠-胶体金免疫层析法可作为水产品中地西泮残留快速检测的有效手段。

Rapid Determination of Diazepam Residue in Aquatic Products by Immunomagnetic Beads-Colloidal Gold Immunochromatography Assay

Aiming at the problem of diazepam residues in aquatic products,a rapid and sensitive detection method of immunomagnetic beads-colloidal gold immunochromatography assay was set up. The immunomagnetic beads prepared by coupling streptavidin magnetic beads with biotinylated diazepam monoclonal antibody were used to rapidly enrich and separate the residues of diazepam in aquatic products,and the colloidal gold immunochromatography assay was used for semi-quantitative detection. The optimized results showed that concentration of colloidal gold labelled antibody,antigen(T-line)and sheep anti-mouse IgG(C-line)were 25 μg/mL,0.6 and 1.5 mg/mL,respectively. The spraying quantity of colloidal gold labelled antibody was 3.0 μL/cm. When diazepam was spiked at the concentration of 1.0,5.0,10.0 and 15.0 μg/kg,the recovery range was from 78.34% to 90.40%,and RSD was from 5.03% to 8.96%. This method had strong specificity and good stability,and the detection could be finished within 25 min. The limit of detection of diazepam residues in aquatic products was 0.5 μg/kg,which was better than the conventional pretreatment method. Furthermore,40 samples were analyzed and the results were consistent with those obtained in GC-MS. It showed that the immunomagnetic beads-colloidal gold immunochromatography assay could be used as an effective method for the rapid determination of diazepam residues in aquatic products.