Development of fluorescence-based receptor binding assays for the detection

Seatox Research, Inc., Wilmington NC

Investigators

Abstract

DESCRIPTION (provided by applicant): Harmful algal blooms (HABs) are becoming frequent occurrences off the coasts of the United States, particularly in coastal waters of New England, the Gulf of Mexico, and the Pacific Ocean. Historically, HABs have been associated with fish kills and marine mammal mortalities; however, their effects on human health and economic loss due to HAB contamination of seafood are becoming more prevalent, with conservative estimates around $82 million annually. The toxic effects of HABs are precipitated by the production of toxins by various species of marine algae and microorganisms, which are then consumed by filter feeding shellfish and finfish. Accumulation in significant quantities can result in toxicity and death in marine animals and humans. Many of the natural toxins produced by marine phytoplankton are heat and acid stable; therefore, cooking contaminated seafood does not eliminate the risk of poisoning. Currently, US state agencies monitor for the presence of toxic phytoplankton and when the cell count reaches a set level, shellfish beds and finfish are tested for the presence of toxin. When toxin levels reach FDA set limits, fishery resources are closed. However, current methods for detection of marine toxins most commonly associated with seafood poisonings in the US have serious drawbacks, including lengthy assay time, high cost, animal usage, low sensitivity and/or sample throughput, or small working ranges. The purpose of this project is to develop a fluorescence based receptor binding assay (FBA) for the detection of marine neurotoxins that cause paralytic shellfish poisoning (saxitoxin) and amnesic shellfish poisoning (domoic acid). These assays could be used as a rapid test alternative to current methods and they have the advantage of lower cost, high sensitivity and lower animal usage. The FBA will be an improvement over the radioligand receptor binding assay (RBA) for these toxins, as the fluorescence platform does not require the use of radioactivity and is thus safer and far less expensive. Using the same techniques as the FBA that we recently developed for brevetoxins and ciguatoxins, fluorophores will be conjugated to saxitoxin and domoic acid and used as the labeled ligand to examine interactions with the toxins' receptors. As RBAs have been used for a variety of sample matrices and been shown to strongly correlate with the mouse bioassay and HPLC results in samples, FBAs could be the next progressive step in the detection of toxins in seafood or coastal water samples thereby protecting human health and aiding in the monitoring of fishery resources.