title: Coupling porous sheathless interface MS with transient-ITP in neutral capillaries for improved sensitivity in glycopeptide analysis.
abstract: IgG antibodies are modulated in their function by the specific structure of the N-glycans attached to their Fc (fragment crystallizable) portions. However, the glycosylation analysis of antigen-specific IgGs is a challenging task as antibody levels to a given antigen only represent a fraction of the total IgG levels. Here, we investigated the use of a transient-ITP (t-ITP)--MS method for highly sensitive IgG1 glycosylation profiling as a complementary method to a high-throughput nano-RPLC-MS method. It was found that t-ITP-CZE using neutrally coated separation capillaries with a large volume injection (37% of capillary volume) and interfaced to MS with a sheathless porous sprayer yielded a 40-fold increase in sensitivity for IgG1 Fc glycopeptide analysis when compared to the conventional strategy. Furthermore, the glycoform profiles found with the t-ITP-CZE strategy were comparable to those from nano-RPLC-MS. In conclusion, the use of the highly sensitive t-ITP-CZE-MS method will provide information on IgG Fc glycosylation for those samples with IgG1 concentrations below the LODs of the conventional method.
authors.affiliation: Biomolecular Mass Spectrometry Unit, Department of Parasitology, Leiden University Medical Center, Leiden, The Netherlands. a.a.m.heemskerk@lumc.nl
authors.forename: Anthonius A M
authors.initials: AA
authors.lastname: Heemskerk
authors.forename: Manfred
authors.initials: M
authors.lastname: Wuhrer
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Busnel
authors.forename: Carolien A M
authors.initials: CA
authors.lastname: Koeleman
authors.forename: Maurice H J
authors.initials: MH
authors.lastname: Selman
authors.forename: Gestur
authors.initials: G
authors.lastname: Vidarsson
authors.forename: Rick
authors.initials: R
authors.lastname: Kapur
authors.forename: Bart
authors.initials: B
authors.lastname: Schoenmaker
authors.forename: Rico J E
authors.initials: RJ
authors.lastname: Derks
authors.forename: André M
authors.initials: AM
authors.lastname: Deelder
authors.forename: Oleg A
authors.initials: OA
authors.lastname: Mayboroda
mesh_terms: D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D006020:Glycopeptides / Q000032:analysis* / Q000097:blood / Q000737:chemistry / Q000302:isolation & purification; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry; D007074:Immunoglobulin G / Q000032:analysis / Q000097:blood / Q000737:chemistry / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation / Q000379:methods*
chemical_list: D006020:Glycopeptides; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G