LC-MS analysis of polyclonal IgGs using IdeS enzymatic proteolysis for oxidation monitoring.
Susceptibility of IgGs to oxidation is a significant issue for intravenous immunoglobulin preparations (IVIG) in liquid solution and raises both safety and efficacy concerns. Here we present an optimized chromatography method coupled to mass spectrometry (MS) to determine the oxidation of Fc/2 fragments derived from polyclonal IgGs after IdeS treatment. Separation of the four IgG subclasses was achieved using a diphenyl column and UV/MS detections were used for quantification and characterization. Several oxidized Fc/2 fragments generated by stress conditions were resolved and oxidized methionines were identified. This procedure can be used to monitor the oxidative status of IVIG preparations during formulation or stability studies.
Analytical Department, LFB Biotechnologies, 3 Avenue des Tropiques, 91958 Courtaboeuf (Les Ulis), France.
Yann
Y
Leblanc
Analytical Department, LFB Biotechnologies, 3 Avenue des Tropiques, 91958 Courtaboeuf (Les Ulis), France.
Marie
M
Romanin
Analytical Department, LFB Biotechnologies, 3 Avenue des Tropiques, 91958 Courtaboeuf (Les Ulis), France.
Nicolas
N
Bihoreau
Analytical Department, LFB Biotechnologies, 3 Avenue des Tropiques, 91958 Courtaboeuf (Les Ulis), France. Electronic address: chevreuxg@lfb.fr.
Guillaume
G
Chevreux
D001426:Bacterial Proteins / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D057056:Cysteine Proteases / Q000378:metabolism; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000302:isolation & purification; D007074:Immunoglobulin G / Q000032:analysis*; D016756:Immunoglobulins, Intravenous / Q000592:standards; D013058:Mass Spectrometry / Q000379:methods; D010084:Oxidation-Reduction; D059748:Proteolysis
D001426:Bacterial Proteins; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D016756:Immunoglobulins, Intravenous; C442815:Mac-1-like protein, Streptococcus; D057056:Cysteine Proteases
IVIG; IdeS; IgG; Mass spectrometry; Oxidation