Safety of milk may be affected by toxic contamination. Aflatoxin B1 (AFB1) is a secondary metabolite produced by some Aspergillus spp. fungi affecting many crops and feed materials. Milk is the only food of animal origin where a significant aflatoxin feed-food carry over may occur. The main AFB1-related compound present in milk is aflatoxin M1 (AFM1), the 4-hydroxylated metabolite of AFB1, classified by the IARC as "group 1 human carcinogen". Regulation No. 1881/2006/EC established the maximum level for AFM1 in raw milk, heat-treated milk and milk for the manufacture of dairy products at 50 ng/kg. Scientific and technological research is called to develop cost- and time-effective field methods that can be transferred to producers for self-control purposes. When validated according to official guidelines,such as those defined in the Commission Regulation 519/2014/EU, rapid methods are expected to complement the consolidated European system for official control that is actually based on sophisticated and expensive laboratory instruments and techniques requiring extensive sample pre-treatment and personnel training. Aim of this work was to evaluate and compare analytical performances of widely applied commercial immunoassays for the detection of AFM1 in raw cow milk, namely lateral flow immunoassay (LFD) and ELISA. Analytical performances such as precision profile, cuf-off, false positive and false negative rates were evaluated for each assay by single laboratory validation performed at AFM1 levels of 0, 25, 50 and 75 ng/kg. Fifty ng/kg was the screening target concentration (STC), that was the concentration of interest for the detection of the mycotoxin in the milk sample. The two assays showed similar performances in terms of cut-off (37.7 ng/kg and 39.4 ng/kg for LFD and ELISA respectively), false suspects rate for blanks (< 0.1% for both assays) and false negative rate for samples containing AFM1 at levels higher than STC (0.3% for both assays). False suspect rate for samples contaminated at 25 ng/kg (50% STC) were 3% and 23% for LFD and ELISA respectively. Similar values were also obtained for the precision at all tested validation levels which was < 19% for the LFD and < 24 %for the ELISA.Furthermore, a satisfactory correlation of the results obtained with the rapid immunoassays and the AOAC Official Method 2000.08 was obtained for the analysis of cow milk samples naturally contaminated at AFM1 levels in the range n.d. - 50 ng/kg. Finally, the extension of the scope of the LFD method to goat and sheep milk was evaluated by applying the experimental design foreseen in the EU regulation. The obtained data showed the applicability of the LFD immunoassay to goat and sheep milk provided that a specific calibration curve was used.
Critical comparison of analytical performances of immunoassay platforms for rapid aflatoxin M1 detection in milk
Michelangelo Pascale;Biancamaria Ciasca;
2018
Abstract
Safety of milk may be affected by toxic contamination. Aflatoxin B1 (AFB1) is a secondary metabolite produced by some Aspergillus spp. fungi affecting many crops and feed materials. Milk is the only food of animal origin where a significant aflatoxin feed-food carry over may occur. The main AFB1-related compound present in milk is aflatoxin M1 (AFM1), the 4-hydroxylated metabolite of AFB1, classified by the IARC as "group 1 human carcinogen". Regulation No. 1881/2006/EC established the maximum level for AFM1 in raw milk, heat-treated milk and milk for the manufacture of dairy products at 50 ng/kg. Scientific and technological research is called to develop cost- and time-effective field methods that can be transferred to producers for self-control purposes. When validated according to official guidelines,such as those defined in the Commission Regulation 519/2014/EU, rapid methods are expected to complement the consolidated European system for official control that is actually based on sophisticated and expensive laboratory instruments and techniques requiring extensive sample pre-treatment and personnel training. Aim of this work was to evaluate and compare analytical performances of widely applied commercial immunoassays for the detection of AFM1 in raw cow milk, namely lateral flow immunoassay (LFD) and ELISA. Analytical performances such as precision profile, cuf-off, false positive and false negative rates were evaluated for each assay by single laboratory validation performed at AFM1 levels of 0, 25, 50 and 75 ng/kg. Fifty ng/kg was the screening target concentration (STC), that was the concentration of interest for the detection of the mycotoxin in the milk sample. The two assays showed similar performances in terms of cut-off (37.7 ng/kg and 39.4 ng/kg for LFD and ELISA respectively), false suspects rate for blanks (< 0.1% for both assays) and false negative rate for samples containing AFM1 at levels higher than STC (0.3% for both assays). False suspect rate for samples contaminated at 25 ng/kg (50% STC) were 3% and 23% for LFD and ELISA respectively. Similar values were also obtained for the precision at all tested validation levels which was < 19% for the LFD and < 24 %for the ELISA.Furthermore, a satisfactory correlation of the results obtained with the rapid immunoassays and the AOAC Official Method 2000.08 was obtained for the analysis of cow milk samples naturally contaminated at AFM1 levels in the range n.d. - 50 ng/kg. Finally, the extension of the scope of the LFD method to goat and sheep milk was evaluated by applying the experimental design foreseen in the EU regulation. The obtained data showed the applicability of the LFD immunoassay to goat and sheep milk provided that a specific calibration curve was used.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.