He Shuang, Liang Demei, Xiong Jincheng, Wang Zile, Zheng Pimiao, Zhang Huixia, Ren Zhenhui, Jiang Haiyang
Department of Veterinary Pharmacology and Toxicology, College of Veterinary Medicine, China Agricultural University, Beijing Key Laboratory of Detection Technology for Animal-Derived Food Safety, Beijing Laboratory for Food Quality and Safety, Beijing 100193, People's Republic of China.
Department of Veterinary Pharmacology and Toxicology, College of Veterinary Medicine, China Agricultural University, Beijing Key Laboratory of Detection Technology for Animal-Derived Food Safety, Beijing Laboratory for Food Quality and Safety, Beijing 100193, People's Republic of China.
J Pharm Biomed Anal. 2022 May 30;214:114719. doi: 10.1016/j.jpba.2022.114719. Epub 2022 Mar 15.
In this study, a sensitive, rapid, homogeneous, and high-throughput fluorescence polarization immunoassay (FPIA) for the rapid screening of eight glucocorticoids (GCs) in beef samples was successfully established. Two tracers including 5-aminofluorescein-labeled dexamethasone (5-AF-DMS) and fluorescein isothiocyanate-labeled dexamethasone (FITC-DMS) were studied to select appropriate antibody-tracer pairs using four previously produced broad-specific monoclonal antibodies. An optimal combination of the antibody 12D9 and the tracer FITC-DMS was selected. Under optimal detection conditions, the half inhibitory concentrations of dexamethasone (DMS), betamethasone (BMS), prednisolone (PNS), hydrocortisone (HCS), beclomethasone (BCMS), cortisone (CS), 6-α-methylprednisone (6-α-MPNS), fludrocortisone acetate (HFCS) were 1.00, 2.17, 3.49, 12.45, 1.20, 5.66, 6.85 and 3.45 ng/mL, respectively. The average recoveries of the proposed method in beef samples ranged from 77.3-91.7% with the coefficient of variation less than 12%. The developed FPIA was time-saving that could be completed within 10 min. The FPIA was applied to beef samples and showed a good correlation with liquid chromatography-tandem mass spectrometry (LC-MS/MS) (R = 0.9894). Thus, the proposed method provides a rapid, reliable, sensitive, and high-throughput screening tool for the simultaneous screening of eight GCs in beef, which shows great potential in the food safety analysis.
在本研究中，成功建立了一种灵敏、快速、均相且高通量的荧光偏振免疫分析法（FPIA），用于牛肉样品中8种糖皮质激素（GCs）的快速筛查。研究了两种示踪剂，即5-氨基荧光素标记的地塞米松（5-AF-DMS）和异硫氰酸荧光素标记的地塞米松（FITC-DMS），使用四种先前制备的广谱特异性单克隆抗体来选择合适的抗体-示踪剂对。选择了抗体12D9和示踪剂FITC-DMS的最佳组合。在最佳检测条件下，地塞米松（DMS）、倍他米松（BMS）、泼尼松龙（PNS）、氢化可的松（HCS）、倍氯米松（BCMS）、可的松（CS）、6-α-甲基泼尼松（6-α-MPNS）、醋酸氟氢可的松（HFCS）的半数抑制浓度分别为 1.00、2.17、3.49、12.45、1.20、5.66、6.85和3.45 ng/mL。所提出方法在牛肉样品中的平均回收率为77.3% - 91.7%，变异系数小于12%。所开发的FPIA省时，可在10分钟内完成。该FPIA应用于牛肉样品，并与液相色谱 - 串联质谱法（LC-MS/MS）显示出良好的相关性（R = 0.9894）。因此，所提出的方法为牛肉中8种GCs的同时筛查提供了一种快速、可靠、灵敏且高通量的筛查工具，在食品安全分析中显示出巨大潜力。
