title: Development of an ELISA for the Detection of Azaspiracids.
abstract: Azaspiracids (AZAs) are a group of biotoxins that cause food poisoning in humans. These toxins are produced by small marine dinoflagellates such as Azadinium spinosum and accumulate in shellfish. Ovine polyclonal antibodies were produced and used to develop an ELISA for quantitating AZAs in shellfish, algal cells, and culture supernatants. Immunizing antigens were prepared from synthetic fragments of the constant region of AZAs, while plate coating antigen was prepared from AZA-1. The ELISA provides a sensitive and rapid analytical method for screening large numbers of samples. It has a working range of 0.45-8.6 ng/mL and a limit of quantitation for total AZAs in whole shellfish at 57 μg/kg, well below the maximum permitted level set by the European Commission. The ELISA has good cross-reactivity to AZA-1-10, -33, and -34 and 37-epi-AZA-1. Naturally contaminated Irish mussels gave similar results whether they were cooked or uncooked, indicating that the ELISA also detects 22-carboxy-AZA metabolites (e.g., AZA-17 and AZA-19). ELISA results showed excellent correlation with LC-MS/MS analysis, both for mussel extract spiked with AZA-1 and for naturally contaminated Irish mussels. The assay is therefore well suited to screening for AZAs in shellfish samples intended for human consumption, as well as for studies on AZA metabolism.
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750 Sentrum, N-0106 Oslo, Norway.
authors.forename: Ingunn A
authors.initials: IA
authors.lastname: Samdal
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750 Sentrum, N-0106 Oslo, Norway.
authors.forename: Kjersti E
authors.initials: KE
authors.lastname: Løvberg
authors.affiliation: AgResearch, Ruakura, East Street, Private Bag 3123, Hamilton, New Zealand.
authors.forename: Lyn R
authors.initials: LR
authors.lastname: Briggs
authors.affiliation: Marine Institute , Rinville, Oranmore, County Galway, Ireland.
authors.forename: Jane
authors.initials: J
authors.lastname: Kilcoyne
authors.affiliation: Department of Chemistry, University of Minnesota-Twin Cities , Minneapolis, Minnesota 55455, United States.
authors.forename: Jianyan
authors.initials: J
authors.lastname: Xu
authors.affiliation: Department of Chemistry and Biochemistry, The Ohio State University , Columbus, Ohio 43210, United States.
authors.forename: Craig J
authors.initials: CJ
authors.lastname: Forsyth
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750 Sentrum, N-0106 Oslo, Norway.
authors.forename: Christopher O
authors.initials: CO
authors.lastname: Miles
mesh_terms: D000818:Animals; D049872:Bivalvia / Q000737:chemistry* / Q000469:parasitology; D004141:Dinoflagellida / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D008387:Marine Toxins / Q000032:analysis* / Q000378:metabolism; D012758:Shellfish / Q000032:analysis* / Q000469:parasitology; D013141:Spiro Compounds / Q000032:analysis* / Q000378:metabolism; D000095702:Polyether Toxins
chemical_list: D008387:Marine Toxins; D013141:Spiro Compounds; C406592:azaspiracid; D000095702:Polyether Toxins
keywords: AZA-1; ELISA; antibody; azaspiracid; immunoassay; mussel; polyclonal; shellfish toxin