title: Trap for MAbs: characterization of intact monoclonal antibodies using reversed-phase HPLC on-line with ion-trap mass spectrometry.
abstract: For the first time, the characterization of intact 150-kDa monoclonal antibodies (MAbs) using a commercially available three-dimensional ion-trap mass spectrometer (IT-MS) is reported. The IT-MS analysis was performed on-line with reversed-phase high performance liquid chromatography (RP-HPLC) on a POROS column using a nontraditional solvent system of acetonitrile, isopropanol, ethanol, and water in formic acid. The operating parameters of the IT-MS were optimized by extending the mass range to m/z 4000 and elevating the tube lens offset voltage value to around -100 V. Mass accuracy better than 300 ppm (+/-40 Da) has been routinely achieved for these macromolecules. Multiple peaks 162 Da apart due to the hexose variants of the monoclonal IgG antibodies were partially resolved in mass spectra. Several commercial and chimeric antibodies have been investigated in this study.
authors.affiliation: Department of Pharmaceutics and Drug Delivery, Amgen Inc., Thousand Oaks, California 91320-1789, USA. johnl@amgen.com
authors.forename: John C
authors.initials: JC
authors.lastname: Le
authors.forename: Pavel V
authors.initials: PV
authors.lastname: Bondarenko
mesh_terms: D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000737:chemistry; D061067:Antibodies, Monoclonal, Humanized; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D007074:Immunoglobulin G / Q000032:analysis; D013058:Mass Spectrometry / Q000379:methods; D008970:Molecular Weight; D012997:Solvents; D000068878:Trastuzumab
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D007074:Immunoglobulin G; D012997:Solvents; D000068878:Trastuzumab
references.citation: Anal Chem. 1995 Oct 15;67(20):3626-32
references.pmid: 8644915
references.citation: Immunol Today. 2000 Aug;21(8):397-402
references.pmid: 10916143
references.citation: J Chromatogr A. 2004 Oct 22;1053(1-2):299-305
references.pmid: 15543996
references.citation: J Chromatogr A. 1995 Sep 29;712(1):177-90
references.pmid: 8556150
references.citation: Bioconjug Chem. 1996 Jan-Feb;7(1):16-22
references.pmid: 8741986
references.citation: Anal Chem. 1996 Jan 1;68(1):1-8
references.pmid: 8779426
references.citation: J Chromatogr B Biomed Sci Appl. 2001 Mar 10;752(2):233-45
references.pmid: 11270864
references.citation: J Mass Spectrom. 2000 Apr;35(4):493-503
references.pmid: 10797646
references.citation: J Chromatogr A. 2001 Aug 24;927(1-2):61-76
references.pmid: 11572399
references.citation: Anal Chem. 1994 Jan 1;66(1):126-33
references.pmid: 8116874
references.citation: J Am Soc Mass Spectrom. 1994 May;5(5):350-8
references.pmid: 24222589
references.citation: Anal Chem. 1995 Oct 15;67(20):3613-25
references.pmid: 8644914
references.citation: Biochemistry. 1997 Dec 2;36(48):14914-23
references.pmid: 9398215
references.citation: Anal Chem. 1994 Mar 1;66(5):585-95
references.pmid: 8154587
references.citation: Biophys J. 2001 Dec;81(6):3503-9
references.pmid: 11721011
references.citation: FEBS Lett. 2000 May 19;473(3):349-57
references.pmid: 10818239
references.citation: Science. 1989 Oct 6;246(4926):64-71
references.pmid: 2675315
references.citation: J Am Soc Mass Spectrom. 1994 Feb;5(2):64-71
references.pmid: 24222517
references.citation: Anal Chem. 2003 Apr 15;75(8):1805-11
references.pmid: 12713037
references.citation: Anal Chem. 1995 Oct 15;67(20):3802-5
references.pmid: 8644926