A New Method for Determination of Alfatoxin M1 in Milk by Ultrasensitive Time-Resolved Fluoroimmunoassay

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, semi-skimmed, 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.