title: A monoclonal antibody-based immunoassay for detecting tetrodotoxin in biological samples.
abstract: Spleen cells from mice hyperimmunized with a keyhole limpet hemocyanin-tetrodotoxin-formaldehyde conjugate were fused with murine P3X63Ag8.653 myeloma cells. A single hybridoma clone was identified that secretes an IgG1,k monoclonal antibody (MAb), designated T20G10, against tetrodotoxin (TTX), with an estimated affinity of 1.2 x 10(8) L/M. Competitive inhibition enzyme immunoassays (CIEIAs) for detecting TTX were developed using this MAb. A direct CIEIA using alkaline phosphatase-labeled MAb detected TTX with sensitivities at IC50 and IC20 of 6-7 ng/ml and 2-3 ng/ml, respectively. The accuracy of the direct CIEIA was comparable with the high-performance liquid chromatography (HPLC) and the mouse bioassay systems, but the direct CIEIA exhibited greater sensitivity. The direct CIEIA was also more cost effective, as it required less sample preparation, a shorter assay time, and reduced investment in equipment than either of the other assay systems.
authors.affiliation: Hawaii Biotechnology Group, Inc., Aiea 96701.
authors.forename: T J
authors.initials: TJ
authors.lastname: Raybould
authors.forename: G S
authors.initials: GS
authors.lastname: Bignami
authors.forename: L K
authors.initials: LK
authors.lastname: Inouye
authors.forename: S B
authors.initials: SB
authors.lastname: Simpson
authors.forename: J B
authors.initials: JB
authors.lastname: Byrnes
authors.forename: P G
authors.initials: PG
authors.lastname: Grothaus
authors.forename: D C
authors.initials: DC
authors.lastname: Vann
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D001681:Biological Assay / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D005069:Evaluation Studies as Topic; D006433:Hemocyanins / Q000276:immunology; D007124:Immunoenzyme Techniques / Q000706:statistics & numerical data; D051379:Mice; D012680:Sensitivity and Specificity; D013779:Tetrodotoxin / Q000032:analysis* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D013779:Tetrodotoxin; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin