Development of an Ultrasensitive and Rapid Fluorescence Polarization Immunoassay for Ochratoxin A in Rice

Ochratoxin A (OTA) is a known food contaminant that affects a wide range of food and agricultural products. The presence of this fungal metabolite in foods poses a threat to human health. Therefore, various detection and quantification methods have been developed to determine its presence in foods. Herein, we describe a rapid and ultrasensitive tracer-based fluorescence polarization immunoassay (FPIA) for the detection of OTA in rice samples. Four fluorescent tracers OTA-fluorescein thiocarbamoyl ethylenediamine (EDF), OTA-fluorescein thiocarbamoyl butane diamine (BDF), OTA-amino-methyl fluorescein (AMF), and OTA-fluorescein thiocarbamoyl hexame (HDF) with fluorescence polarization values (delta FP = FPbind-FPfree) of 5, 100, 207, and 80 mP, respectively, were synthesized. The tracer with the highest delta FP value (OTA-AMF) was selected and further optimized for the development of an ultrasensitive FPIA with a detection range of 0.03-0.78 ng/mL. A mean recovery of 70.0% to 110.0% was obtained from spiked rice samples with a relative standard deviation of equal to or less than 20%. Good correlations (r(2) = 0.9966) were observed between OTA levels in contaminated rice samples obtained by the FPIA method and high-performance liquid chromatography (HPLC) as a reference method. The rapidity of the method was confirmed by analyzing ten rice samples that were analyzed within 25 min, on average. The sensitivity, accuracy, and rapidity of the method show that it is suitable for screening and quantification of OTA in food samples without the cumbersome pre-analytical steps required in other mycotoxin detection methods.