Competitive ELISA: An Accurate, Quick and Effective Tool to Monitor Brevetoxins in Environmental and Biological Sample

Abstract

A competitive Enzyme-Linked Immuno-Sorbent Assay (competitive ELISA) has been developed for analyzing brevetoxins (PbTxs). Antibodies to brevetoxins were used in combination with a multi-step signal amplification procedure for the detection of toxins. This procedure minimizes non-specific signals and background noise often observed in complex matrices. Therefore, analysis can be performed with various samples (seawater, air filter, mammalian body fluids, shellfish, etc.) without the need for extensive extraction and/or purification steps. Brevetoxin analysis in liquid samples like seawater, urine and serum can be performed without pretreatment, dilution or purification. The limit of quantification of PbTxs is 2 ng mL^-1 in any of the liquid sample matrices tested. For shellfish monitoring, analyses are performed after homogenization of shellfish meat (5 g) with brevetoxin-ELISA buffer (200 mL) and can be performed on tissue from a single mollusk as well as on a pool of shellfish meat. Comparative quantification of PbTxs achieved in buffer, seawater, mammalian body fluid and shellfish homogenate spiked with equal amounts of toxin (10 ng mL^-1 sample) varied by no more than 5%. These data suggest that the matrix composition of the sample does not affect the performance of the assay. Because this assay is not affected by matrix composition and can be performed in shellfish homogenate, this procedure can be used to prevent or diagnose human exposure to PbTxs and has the potential to replace the currently used mouse bioassay for monitoring PbTxs in shellfish.