title: Use of T-2 toxin-immobilized amine-activated beads as an efficient affinity purification matrix for the isolation of specific IgY.
abstract: An affinity purification method that isolates T-2 toxin-specific IgY utilizing a T-2-toxin-immobilized column was developed. The T-2 toxin was covalently coupled via a carbonyldiimidazole-activated hydroxyl functional group to amine-activated sepharose beads. The affinity-purified IgY was characterized by gel electrophoresis, fast protein liquid chromatography, enzyme-linked immunosorbant assay, surface plasmon resonance and mass spectrometry. A competitive inhibition ELISA (CI-ELISA) was performed using affinity-purified IgY with a T-2 toxin detection sensitivity of 30 ng/mL, which falls within the maximum permissible limit of 100 ng/mL. The cross reactivity of IgY towards deoxynivalenol, zearalenone, fumonisin B1 and HT-2 was significantly reduced after affinity purification. A surface plasmon resonance (SPR)-based inhibition assay was also applied for quantitative determination of T-2 toxin in spiked wheat samples. The results obtained indicate the feasibility of utilizing this IgY-based assay for the detection of T-2 toxin in food samples.
authors.affiliation: Applied Biochemistry Group (ABG), School of Biotechnology, Dublin City University, Dublin 9, Ireland.
authors.forename: Soujanya Ratna
authors.initials: SR
authors.lastname: Edupuganti
authors.forename: Om Prakash
authors.initials: OP
authors.lastname: Edupuganti
authors.forename: Richard
authors.initials: R
authors.lastname: O'Kennedy
authors.forename: Eric
authors.initials: E
authors.lastname: Defrancq
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Boullanger
mesh_terms: D000818:Animals; D002645:Chickens; D002846:Chromatography, Affinity / Q000295:instrumentation* / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D007136:Immunoglobulins / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D013058:Mass Spectrometry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance; D013605:T-2 Toxin / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D014908:Triticum / Q000737:chemistry
chemical_list: C016512:IgY; D007136:Immunoglobulins; D013605:T-2 Toxin