|Name||Miss Amanda Bartley|
|Organization or Institution||Florida International University|
Aptamer-Based Assay for Detection of Ochratoxin A in Wine
Amanda Bartley; John P. Berry
Florida International University
Ochratoxin A (OTA) is a potent mycotoxin found in a wide range of agricultural products. Originating from fungal genera of Aspergillus and Penicillium, pathologies associated with OTA include mitochondrial damage and renal disease (e.g., “Balkan Endemic Nephropathy”), and it is considered a class 2B possible carcinogen by the International Agency for Research on Cancer. The standard methods for OTA analysis currently rely on the use of high-performance liquid chromatography (HPLC) coupled to either fluorescence detection or mass spectrometry, and typically require substantial sample preparation and/or clean up steps. Toward a high-throughput analysis of OTA, we propose a method based on highly specific aptamer-based detection. The proposed aptamer-based assay utilizes a single-stranded DNA aptamer specific to OTA, modified with a fluorophore, coupled to a complementary sequence modified with a FRET-based quencher which dissociates in the presence of the target toxin. The method has been developed using two fluorophore/quencher pairs, namely FAM/TAMRA and CY5/IABRQ, and multiple methods (e.g., biotin/streptavidin, SMCC/polylysine) for immobilization of the fluorophore-labeled aptamer within a 96-well plate format. The developed assay will be evaluated and validated using wine as a representative agricultural source of OTA. The proposed high-throughput assay would provide a considerable improvement in the detection of OTA as a human health concern.