Microsphere-based immunoassay for the detection of azaspiracids.
Azaspiracids (AZAs) are a group of lipophilic toxins discovered in mussels from Ireland in 1995 following a human poisoning incident. Nowadays the regulatory limit for AZAs in many countries is set at 160 μg of azaspiracid equivalents per kilogram of shellfish meat. In this work a microsphere-based immunoassay has been developed for the detection of AZAs using a Luminex system. This method is based on the competition between AZA-2 immobilized onto the surface of microspheres and free AZAs for the interaction with a monoclonal anti-azaspiracid antibody (mAb 8F4). In this inhibition immunoassay the amount of mAb 8F4 bound to AZA-2 microspheres was quantified using a phycoerythrin-labeled anti-mouse antibody, and the fluorescence was measured with a Luminex analyzer. Simple acetate/methanol or methanol extractions yielded final extracts with no matrix interferences and adequate recovery rates of 86.5 and 75.8%, respectively. In summary, this work presents a sensitive and easily performed screening method capable of detecting AZAs at concentrations below the range of the European regulatory limit using a microsphere/flow cytometry system.
Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002 Lugo, Spain.
Laura P
LP
Rodríguez
Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002 Lugo, Spain. Electronic address: natalia.vilarino@usc.es.
Natalia
N
Vilariño
Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002 Lugo, Spain.
M Carmen
MC
Louzao
Department of Chemistry, The Scripps Research Institute, La Jolla, CA 92037, USA; Worm Institute for Research and Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA.
Tobin J
TJ
Dickerson
Department of Chemistry, The Scripps Research Institute, La Jolla, CA 92037, USA; Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA; Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, CA 92093, USA; Department of Chemistry, BioScience Research Collaborative, Rice University, Houston, TX 77030, USA.
K C
KC
Nicolaou
Department of Chemistry, The Scripps Research Institute, La Jolla, CA 92037, USA; Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA; Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, CA 92093, USA.
Michael O
MO
Frederick
Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002 Lugo, Spain. Electronic address: luis.botana@usc.es.
Luis M
LM
Botana
D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D005470:Fluorometry; D007118:Immunoassay / Q000379:methods*; D008387:Marine Toxins / Q000032:analysis*; D008863:Microspheres; D013141:Spiro Compounds / Q000032:analysis*; D013997:Time Factors; D000095702:Polyether Toxins
D000911:Antibodies, Monoclonal; D008387:Marine Toxins; D013141:Spiro Compounds; C406592:azaspiracid; D000095702:Polyether Toxins
Anti-azaspiracid antibody; Azaspiracid poisoning; Flow fluorimetry system