A competitive fluorescent aptasensor for okadaic acid detection assisted by rolling circle amplification

Okadaic acid (OA) is a low molecular weight marine toxin from shellfish which causes diarrheic shellfish poisoning (DSP). Due to its frequent occurrence, OA has become a serious threat to human health and seafood industry. The authors describe a competitive fluorophore-linked aptamer assay for OA that is based on rolling circle amplification (RCA). It consists of the following steps: (a) The wells of a microplate are modified by fixing the OA aptamer on their surface; (b) The aptamer is hybridized with an aptamer-complementary sequence-primer complex; (c) the RCA reaction is performed; (d) the FAM labelled signal probe is added. OA competes with the detection probe for the immobilized aptamer. After the competitive reaction has occurred, the supernatants containing released detection probes are removed and then read with a microplate reader. This method, unlike in competitive assays where the signals negatively correlate with OA concentrations, has a positive correlation between fluorescence intensity and OA concentration. The optimized assay has a lower detection limit (1 pg·mL⁻¹) and a wider linear range (from 1 pg·mL⁻¹ to 100 ng·mL⁻¹) owning to signal amplification via RCA. It also is highly specific, repeatable, has good recovery and can be used to detect OA in seafood. .