title: Development of a mass spectrometry immunoassay for unambiguous detection of egg allergen traces in wines.
abstract: A mass spectrometry immunoassay (MSIA) specifically designed for the detection of egg allergens in wines is described. MSIA is based on an immunoaffinity enrichment procedure combined with targeted MS/MS detection of selected egg peptide markers. Polyclonal antibodies raised against native ovalbumin, chosen as the target protein tracing for egg powder, were immobilized onto low backpressure monolithic MSIA customized disposable tips. Ovalbumin-free wine samples were fortified with standard protein at different concentrations in the low microgram-per-milliliter range. A simple protocol was devised consisting of a 1:4 dilution of the wine sample with a basic solution for pH adjustment, followed by a semi-automated purification/enrichment step on MSIA customized disposable tips fitted on a multichannel electronic pipette. Among the main figures of merit, LOD and LOQ values as low as 0.01 and 0.03 μg/mL, respectively, and within-day precision of 18% should be noticed. Noteworthy, the developed assay outperformed current MS-based methods for the detection of allergenic protein in wine matrices, thanks to the immunoaffinity enrichment. In addition, compared to other immunoassays, the present approach boasts the unquestionable advantage of providing an unambiguous identification of the target protein by simultaneous detection of three unique peptide markers each giving three specific MS/MS transitions.
authors.affiliation: Institute of Sciences of Food Production, National Research Council (ISPA-CNR), Via G. Amendola 122/O, 70126, Bari, Italy.
authors.forename: Rosa
authors.initials: R
authors.lastname: Pilolli
authors.affiliation: Thermo Fisher Scientific, Tempe, AZ, 85284, USA.
authors.forename: Ravindra
authors.initials: R
authors.lastname: Chaudhari
authors.affiliation: Dipartimento di Chimica, Università degli Studi di Bari Aldo Moro, Via Orabona, 4, 70126, Bari, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Palmisano
authors.affiliation: Institute of Sciences of Food Production, National Research Council (ISPA-CNR), Via G. Amendola 122/O, 70126, Bari, Italy. linda.monaci@ispa.cnr.it.
authors.forename: Linda
authors.initials: L
authors.lastname: Monaci
mesh_terms: D000485:Allergens / Q000032:analysis*; D000818:Animals; D002645:Chickens; D004531:Eggs / Q000032:analysis; D005506:Food Contamination / Q000032:analysis*; D007118:Immunoassay / Q000379:methods*; D057230:Limit of Detection; D010047:Ovalbumin / Q000032:analysis*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014920:Wine / Q000032:analysis*
chemical_list: D000485:Allergens; D010047:Ovalbumin
keywords: Egg allergen; Immunoassay; MSIA; Mass spectrometry; Ovalbumin; Wine