孔雀石绿的磁性免疫层析快速检测方法研究

Study on the rapid detection method of malachite green by magnetic immunochromatography

开发了一种适用于现场快速检测孔雀石绿(MG)的免疫层析试纸条,在超顺磁性纳米微球上偶联MG单克隆抗体作为检测探针,分别将孔雀石绿完全抗原(MG-BSA)和羊抗鼠IgG喷涂于NC膜的T线和C线。结果发现,T线最佳喷涂量为0.25 mg/mL,抗体最佳偶联量为20μg,构建的试纸条可在25 min内实现养殖用水及鱼肉组织的定性与定量分析,检出限为1μg/mL,定量限分别为0.60,0.97 ng/mL,对无色孔雀石绿无交叉反应,批内、批间相对标准偏差均<12%;通过与HPLC和市售胶体金试剂盒对比发现,水样的回收率在82.0%~117.4%,灵敏度高于市售胶体金试剂盒。该孔雀石绿磁性免疫层析条适用于现场快速检测鱼肉或水体中的残留孔雀石绿。

This study aims to develop an immunochromatographic test strip for rapid on-site detection of malachite green(MG).Superparamagnetic nanospheres were coupled with MG monoclonal antibodies as detection probes,and MG-BSA and goat anti-mouse IgG were sprayed on the T line and C line of the NC membrane,respectively.The optimal concentration of MG-BSA sprayed on T-line and the coupling amount of antibody were 0.25 mg/mL and 20μg,respectively.The magnetic immunochromatographic test strip can realize the qualitative and quantitative analysis of MG in aquaculture water and fish tissue within 25 min.The limit of detection was 1μg/mL,the limits of quantitation were 0.60 and 0.97 ng/mL for aquaculture water and fish tissue,respectively.There was no cross-reaction to leucomalachite green,and the RSDs of the intra-assay and inter-assay were both<12%.By comparing with HPLC and commercially colloidal gold kits,the recoveries in water samples were 76.2%-116%,which had good correlation and higher sensitivity than colloidal gold kits.The malachite green magnetic immunochromatographic strip is suitable for rapid detection of residual malachite green in fish or water on site.