An immunomagnetic-bead-based enzyme-linked immunosorbent assay for sensitive quantification of fumonisin B1

•Rapid and sensitive immunomagnetic-beads based ELISA (IMB-ELISA) was developed.•The detection limit (0.24 ng/mL) was better than the conventional ELISA.•As a new strategy for detection, the IMB-ELISA was accurate, sensitive, and quick.

A sensitive enzyme-linked immunosorbent assay (ELISA) based on immunomagnetic beads (IMB-ELISA) was established using a magnetic-bead signal-enrichment system. The immunomagnetic beads were coated with polyclonal antibody directed against keyhole limpet hemocyanin (KLH), which were then coupled with a KLH-fumonisin B1 (FB1) conjugate. Anti-FB1 monoclonal antibody and sample extract were mixed and added to the immunomagnetic-bead solution. After the addition of horseradish peroxidase (HRP)-labeled goat anti-mouse antibody and the substrate solution, stop solution was added and the optical density of the reaction mixture was determined. To improve the performance of this method, the dilution of the immunomagnetic beads, the concentrations of the monoclonal antibody and HRP-labeled goat anti-mouse antibody, and the incubation time for the competition reaction were optimized. Based on the optimum conditions, the regression equation for this IMB-ELISA in quantifying FB1 was y = −0.3538x + 0.703 (R2 = 0.9988). The detection limit and IC50 were 0.24 ng/mL and 3.17 ng/mL, respectively. The working range was 0.54-26.3 ng/mL. The recovery rates were 80.4-114.7%, when the spiked concentrations ranged from 19.5 to 156.3 μg/kg. This IMB-ELISA is accurate and more sensitive and less time-consuming than the conventional ELISA.

Graphical abstractThe immunomagnetic beads were coated with anti-keyhole limpet hemocyanin (KLH) polyclonal antibody, which then coupled with KLH-fumonisin B1 (FB1) conjugation. The mixture of anti-FB1 monoclonal antibody and sample extractions was added into the immunomagnetic-beads solution. After adding the horseradish peroxidase (HRP)-labeled goat anti-mouse antibody and the substrate solution, the optical density of the reaction mixture was determined after adding stop solution.Download full-size image