title: Detection of Tetrodotoxins in Puffer Fish by a Self-Assembled Monolayer-Based Immunoassay and Comparison with Surface Plasmon Resonance, LC-MS/MS, and Mouse Bioassay.
abstract: The increasing occurrence of puffer fish containing tetrodotoxin (TTX) in the Mediterranean could represent a major food safety risk for European consumers and threaten the fishing industry. The work presented herein describes the development of a new enzyme linked immunosorbent assay (mELISA) based on the immobilization of TTX through dithiol monolayers self-assembled on maleimide plates, which provides an ordered and oriented antigen immobilization and favors the antigen-antibody affinity interaction. The mELISA was found to have a limit of detection (LOD) of TTX of 0.23 mg/kg of puffer fish matrix. The mELISA and a surface plasmon resonance (SPR) immunosensor previously developed were employed to establish the cross-reactivity factors (CRFs) of 5,6,11-trideoxy-TTX, 5,11-deoxy-TTX, 11-nor-TTX-6-ol, and 5,6,11-trideoxy-4-anhydro-TTX, as well as to determine TTX equivalent contents in puffer fish samples. Results obtained by both immunochemical tools were correlated (R(2) = 0.977). The puffer fish samples were also analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS), and the corresponding CRFs were applied to the individual TTX contents. Results provided by the immunochemical tools, when compared with those obtained by LC-MS/MS, showed a good degree of correlation (R(2) = 0.991 and 0.979 for mELISA and SPR, respectively). The mouse bioassay (MBA) slightly overestimated the CRF adjusted TTX content of samples when compared with the data obtained from the other techniques. The mELISA has been demonstrated to be fit for the purpose for screening samples in monitoring programs and in research activities.
authors.affiliation: IRTA , Carretera Poble Nou km 5.5, 43540 Sant Carles de la Ràpita, Tarragona, Spain.
authors.forename: Laia
authors.initials: L
authors.lastname: Reverté
authors.affiliation: IRTA , Carretera Poble Nou km 5.5, 43540 Sant Carles de la Ràpita, Tarragona, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: de la Iglesia
authors.affiliation: IRTA , Carretera Poble Nou km 5.5, 43540 Sant Carles de la Ràpita, Tarragona, Spain.
authors.forename: Vanessa
authors.initials: V
authors.lastname: del Río
authors.affiliation: Institute for Global Food Security, School of Biological Sciences, Queen's University , Stranmillis Road, Belfast BT9 5AG, Northern Ireland.
authors.forename: Katrina
authors.initials: K
authors.lastname: Campbell
authors.affiliation: Institute for Global Food Security, School of Biological Sciences, Queen's University , Stranmillis Road, Belfast BT9 5AG, Northern Ireland.
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Elliott
authors.affiliation: Division of Bacteriology, Osaka Prefectural Institute of Public Health , 3-69, Nakamichi 1-chome, Higashinari-ku, Osaka 537-0025, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Kawatsu
authors.affiliation: National Reference Laboratory on Marine Biotoxins, Ministry of Rural Development and Food , 54627 Thessaloniki, Greece.
authors.forename: Panagiota
authors.initials: P
authors.lastname: Katikou
authors.affiliation: IRTA , Carretera Poble Nou km 5.5, 43540 Sant Carles de la Ràpita, Tarragona, Spain.
authors.forename: Jorge
authors.initials: J
authors.lastname: Diogène
authors.affiliation: IRTA , Carretera Poble Nou km 5.5, 43540 Sant Carles de la Ràpita, Tarragona, Spain.
authors.forename: Mònica
authors.initials: M
authors.lastname: Campàs
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods*; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D057230:Limit of Detection; D051379:Mice; D020349:Surface Plasmon Resonance / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D023721:Tetraodontiformes; D013779:Tetrodotoxin / Q000032:analysis*
chemical_list: D013779:Tetrodotoxin