Polyclonal antibodies to domoic acid, and their use in immunoassays for domoic acid in sea water and shellfish

Abstract

Ovine antibodies raised against conjugates linked through the secondary amino group of domoic acid (1) were used, together with activated-ester-derived conjugates of domoic acid (DA) as the plate coater, to develop a robust indirect competitive enzyme-linked immunosorbent assay (cELISA) for DA in shellfish and seawater. The ELISA was used to analyze shellfish samples for DA, and was compatible with several extraction procedures. The ELISA had a detection limit below 0.01 ng ml(-1), a limit of quantitation (LOQ) of 0.15 ng ml(-1) and a working range of 0.15-15 ng ml(-1) DA. The LOQ is equivalent to 38 ng g(-1) DA in shellfish flesh, assuming a 250-fold dilution during extraction. This is more than 500 times lower than the maximum permitted level (20 microg g(-1) flesh). The ELISA is designed for use alongside regulatory analyses, and, following formal validation, should be available for pre-screening of regulatory shellfish flesh samples. The ELISA was also shown to be appropriate for analysis of DA in algal cultures and in samples of seawater, and thus has the potential to provide early warning of developing algal blooms.

Distribution, Abundance and Domoic Acid Analysis of the Toxic Diatom Genus Pseudo-nitzschia from the Chesapeake Bay

“…via epifluorescent microscopy were processed for domoic acid via ELISA. DA was quantified via an ASP cELISA test kit (Biosense Laboratories) capable of processing a maximum of 36 samples per kit (Bergen, Norway; Garthwaite et al 1998). Each assay has a limit of quantitation (LOQ) and a limit of detection (LOD=1/3 LOQ).…”

Remote sampling of harmful algal blooms: A case study on the Washington State coast

“…We prepared DA immunogen by linking fetuin (FET), a carrier protein, to the secondary amino group of the DA molecule (Garthwaite et al, 1999). Rabbits were immunized subcutaneously with DA amine-linked to a FET conjugate suspended in phosphatebuffered saline (PBS) mixed 1:1 with Freund's complete adjuvant (R&R Rabbitry Research and Development, Stanwood, WA).…”

Development of Polyclonal Antibodies Against Domoic Acid for Their Use in Electrochemical Biosensors

“…Analysis of DA is currently performed by mouse bioassay, [6][7][8] HPLC, [9,10] thin-layer chromatography, [11] and immunochemical methods. [12][13][14] The ideal assay for detection of marine toxins should be simple, cost effective, highly sensitive and specific. For these reasons, methods involving immunological analyses have become popular in recent years.…”