软骨藻酸胶体金免疫层析检测试纸条的研制

Development of Colloidal Gold Immunochromatographic Strip for Rapid Detection of Domoic Acid

制备软骨藻酸(DA)胶体金免疫层析快速检测试纸条,建立一种快速检测贝类食品中软骨藻酸含量的检测方法.采用柠檬酸三钠还原氯金酸法制备颗粒直径20 nm的胶体金;电镜鉴定后,确定胶体金标记软骨藻酸单克隆抗体的最佳条件并制备金标抗体;金标抗体喷涂在玻璃纤维垫上制成胶体金结合垫;软骨藻酸与牛血清蛋白(BSA)的偶联物构成的包被抗原(DA-BSA)和羊抗鼠二抗分别包被在硝酸纤维素膜(NC膜)上构成检测线和控制线;组装成胶体金免疫层析试纸条并进行灵敏度测试.结果显示,胶体金试纸条检测灵敏度为20 ng/mL,检测过程可在15 min内完成.该方法灵敏度高、简单快速,无需特殊仪器设备,适用于广大基层单位的现场和批量检测,有着良好的商业开发应用前景.

The colloidal gold immunochromatographic test strip was developed in order to establish a rapid detection assay of Domoic acid（DA） content in marine shellfish.The colloidal gold with particle diameter 20 nm was obtained by reducing gold chloride with sodium citrate.After identification by electron micrograph,optimum conditions for labeling were determined and colloidal gold was labeled by DA monoclonal antibody.The gold-labeled antibody was coated on some chosen glass fiber and dried.The coating antigen（DA-BSA） and Goat anti Mouse IgG were spotted respectively on a piece of nitrate fiber membrane as test line and control line.Finally the test strips were constructed and the detection sensitivity was measured.The results showed that,the detection limit of colloidal gold immunochromatographic test strip was 20 ng/mL and the whole analysis process could be completed within 15 min.The method established is sensitive and the procedure of determination is simple and quick without special equipment.The colloidal gold immunochromatographic test strip could be widely used for batch detection of domoic acid in shellfish on site and has great prospect for commercial development.