Nanobody-based magnetic chemiluminescence immunoassay for one-pot detection of ochratoxin A

Ochratoxin A (OTA) contamination seriously threatens food safety and human health and requires sensitive and rapid tools for monitoring. In this study, a convenient enzyme-linked immunosorbent assay based on Avi-labeled nanobody Nb-2G/streptavidin-alkaline phosphatase and magnetic beads (MBS-ELISA) was established for the sensitive detection of OTA, which could be used for one-pot detection without immobilization. After optimization, the 50% inhibitory concentration (IC50) and the lowest limit of detection value of the MBS-ELISA was 1.17 ng/mL and 0.07 ng/mL and the linear range was 248.8 pg/mL-5.28 ng/mL, respectively, which accords with state criteria for food safety. The developed one-step MBS-ELISA was almost 20-times more sensitive than the classic BA-ELISA and could generate results within 15 min, which was significantly less than the classic BA-ELISA at approximately 3 h. The MBS-ELISA indicated good recovery (86.4-114.3%) in spiked sorghum, buckwheat, and mung bean. Thus, MBS-ELISA represents a very promising strategy for the simple, rapid, and accurate detection of OTA and other toxic and hazardous contaminants.