A New Method for Determination of Alfatoxin M1 in Milk by Ultrasensitive Time-Resolved Fluoroimmunoassay
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A New Method for Determination of Alfatoxin M1 in Milk by Ultrasensitive Time-Resolved Fluoroimmunoassay Mingming Guo 1 & Bin Zhou 1 & Zijian Huang 2 & Chuncheng Zhao 3 & Jue Zhang 1 & Biao Huang 1
Received: 6 November 2016 / Accepted: 12 February 2017 # Springer Science+Business Media New York 2017
Abstract A competitive indirect time-resolved fluoroimmunoassay (TRFIA) for detecting aflatoxin M1(AFM1) contamination in milk was developed, by using aflatoxin M1-bovineserum albumin conjugate, anti-AFM1 antibody, and Eu-labeled goat anti-rabbit antibody. To improve the sensitivity of the assay, the concentrations of the coating antigen and anti-AFM1 antibody were varied to optimize the condition of the immunological assay. The limit of detection values, limit of quantification values, and dynamic working range were 0.006, 0.022, and 0.022–1.334 μg/kg, respectively. Values of recovery within and between assays were 88.0–116.0% and 92.69– 108.63%. The method was applicable for the full-cream, semiskimmed, skimmed, and raw milk as well. Values of repeatability (intra-laboratory variability) and reproducibility (inter-laboratory variability) were 1.2–4.5% and 0.8–5.0%, respectively. The results of using AFM1-TRFIA to analyze samples of 23 brands of milk that were purchased in Wuxi revealed that AFM1 was absent from all studied samples. This study suggests that the novel method is a simple, sensitive, specific, reproducible, economic, and adequate method for screening large quantities of samples and has good prospects of application. Keywords Time-resolved fluoroimmunoassay . AFM1 . Europium . Milk * Biao Huang [email protected] 1
Key Laboratory of Nuclear Medicine, Ministry of Health/Jiangsu Key Laboratory of Molecular Nuclear Medicine, Jiangsu Institute of Nuclear Medicine, Wuxi 214063, China
2
Department of Biological Sciences, Xi’an Jiaotong-liverpool University, Suzhou 215123, China
3
Beosen(Jiangsu) Food Safety Technology Co. Ltd, Wuxi 214000, China
Introduction Aflatoxins (AFs) are a class of highly toxic and carcinogenic mycotoxins produced primarily by Aspergillus flavus, Aspergillus parasticus, and Aspergillus nomius (Creppy 2002). These toxins are found worldwide in stage of crop growth, harvest, storage, and processing (Bhat et al. 2010; Caloni et al. 2006; Jiang et al. 2005). AFB1 is the most significant one among all subtypes. By the action of P450 cytochrome enzyme, AFM1 is the hydroxylate metabolite of AFB1 in mammary glands that consume AFB1-contaminated diets (Fallah 2010). It has been classified as class 1 human carcinogen by the International Agency for Research on Cancer (IARC 2002), associating with various of toxicity, including carcinogenesis, teratogenesis, and mutagenesis (Corcuera et al. 2011; Roda et al. 2010). To protect public health, most countries define the maximum acceptable levels of AFM1 in milk and dairy products that vary from 0.05 μg/kg in EU (European Commission 2006) to 0.5 μg/kg in China and in the USA. Many methods have been established for AFM1 detection previously. Hig
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