胶体金免疫层析法快速检测配方羊奶粉中的牛β-乳球蛋白

Colloidal Gold Immunochromatographic Assay for Rapid Detection of Bovine β-lactoglobulin in Goat Milk Formulas

建立了一种可快速检测配方羊奶粉中牛β-乳球蛋白(β-lactoglobulin,β-lg)的胶体金免疫层析检测方法。通过杂交瘤技术制备β-lg单克隆抗体(monoclonal antibody,mAb),半抑制浓度(50%inhibiting concentration,IC_(50))为5.87μg/mL。将胶体金标记的β-lg mAb包被于金标垫,β-lg和山羊抗小鼠IgG标记于硝酸纤维素膜(nitrocellulose membrane,NC膜)分别作为检测线(T线)和质控线(C线),开发了可检测β-lg的免疫层析试纸条。该试纸条对β-lg的检测限(limit of detection,LOD)值为50μg/mL,与其他基质成分均未产生有效交叉反应,对全脂山羊乳粉中掺杂脱脂牛奶粉(nonfat skim milk,NFSM)、脱盐乳清粉(desalted whey powder,DWP)和乳清蛋白粉(whey protein powder,WPP)的LOD值分别为5%、5%和0.1%。运用该方法对9个市售配方羊奶粉进行分析,检测结果与商品化酶联免疫吸附测定(enzyme linked immunosorbent assay,ELISA)试剂盒一致。该方法前处理快速简单,5 min即可裸眼判定结果,可用于配方羊奶粉商品的现场快速检测。

The aim of this study was to develop a colloidal gold-based immunochromatographic assay for rapid detection bovine β- lactoglobulin （β-lg） in commercial goat milk formulas. Firstly, an anti-β-lg monoclonal antibody （ mAb ） was prepared using the hybridoma technique with 50% inhibitory concentration（ IC50 ）value of 5.87 μg/mL.To develop an immunochromatographie test strip for detection β-lg, gold labed β- lg mAbs were sprayed on gold conjugation pad, and nitrocellulose membrane was lined with β- lg and goat anti- mouse immunoglobulins as test line （ T- line ） and control line （ C- line ）. The limit of detection （LOD） value of the test strip was 50 μg/mL for β-lg, and no effective cross-reactivities with other matrix ingredients were observed.The LOD of adulteration of whole goat milk powder with cow＇ s nonfat skim milk （ NFSM ） , desahed whey powder （DWP） and whey protein powder （WPP） were 5 % , 5 % and 0.1% , respectively. Furthermore, 9 commercial goat milk formulas were analyzed by this assay, and the resuhs were in accordance with that obtained by commercial enzyme linked immunosorbent assay（ELISA） kit. The assay includes a rapid and simple treatment of sample, could be finished in 5 min, which could be evaluated by naked-eye, was qualified for on-site monitoring goat formula milk adulteration.