Enzyme-linked immunosorbent assay (ELISA) based on superparamagnetic nanoparticles for aflatoxin M1 detection.

Five different clones of antibodies developed against the aflatoxin M(1) were investigated by using the classical indirect and direct competitive Enzyme-Linked Immunosorbent Assay (ELISA) formats, and also the direct competitive ELISA based on the use of the superparamagnetic nanoparticles. The purpose of this study was to assess if not so friendly time classical ELISA procedures can be further improved, by reducing the coating, blocking and competition time. Here we showed that a complete dc-ELISA (coating, blocking and competition step) based on the use of superparamagnetic nanoparticles can be performed in basically 40 min, if coating step (20 min) should be taken into account. Moreover, the standard analytical characteristics of the proposed method fulfil the requirements for detecting AFM(1) in milk, in a wide linear working range (4-250 ng/L). The IC(50) value is 15 ng/L. The matrix effect and the recovery rate were assessed, using the European Reference Material (BD282, zero level of AFM(1)), showing an excellent percentage of recovery, close to 100%.