The NJ Mass Spectrometry Discussion Group presents the June 2013 Meeting on Tuesday, June 18, 2013 at Mirabelle’s Room, DoubleTree by Hilton Somerset, 200 Atrium Drive, Somerset, New Jersey 08873.
Sponsored by Thermo.
Attendance is free of charge, compliments of our sponsor!
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I. Jonathan Josephs, Ph.D
Principal Scientist, Bristol-Myers Squibb
“Creation of Rich MSn Data for Structure Elucidation in Quan/Qual Analysis: Applications of Combined HCD/CID and New Isolation Techniques”
II. Dr. Jie Qian
“Orbitrap Analysis of Native Intact Monoclonal Antibody”
- 5:30 pm Social and registration
6:15 pm Complimentary dinner
7:00 pm Welcome and opening remarks
7:05 pm Jonathan Josephs
8:00 pm Jie Qian
8:55 pm Closing remarks
Abstract for Talk II:
Introduction — Because monoclonal antibodies (mAbs) are increasingly an important line of therapeutics for the biopharmaceutical industry, the demand to better understand the biochemical and biophysical properties of mAbs has become critical. Recent developments in native protein mass spectrometry (MS) have clearly shown its distinctive power for characterization of intact proteins and protein conjugates. The mass spectrometry–based study of intact mAbs in native-like states provides a wealth of information to interpret its biological features. This is the first reported use of a commercial Orbitrap mass analyzer for measuring a native intact mAb protein. A monoclonal IgG1 kappa antibody, composed of two heavy chains of 451 amino acids and two light chains of 213 amino acids was characterized in this study.
Methods — The mAb was analyzed using normal flow liquid chromatography (LC)-MS method. The intact mAb protein was eluted from a size exclusion column under non-denaturing and non-reducing conditions and then directly detected by a standard stock bench-top Orbitrap mass spectrometer under full scan MS mode. The AGC target was set at 3e6 with resolution at 17,500. The MS spectrum was then analyzed with protein deconvolution software (Protein Deconvolution 2.0) that utilizes the ReSpect algorithm for molecular mass determination.
Results — To monitor the IgG1 kappa in native-like state, the protein was dissolved under non-denatured and non-reduced conditions. The protein was then chromatographically eluted from a size exclusion column and directly analyzed online using an Orbitrap mass spectrometer with high mass range. The mass spectra from LC-MS analyses of the native intact antibody demonstrate a clean distribution of the multiply charged envelope at mass range above 5000 m/z. Individual charge states in the raw mass spectrum for the native antibody shows major glycosylation species. Results from this study demonstrate an efficient and potentially high through-put workflow for antibody measurement in native-like states. This protocol, using a combination of normal flow size exclusion LC and a HR/AM bench-top Orbitrap mass spectrometer is expected to provide important data for mAb studies.
Novel Aspect — Native Intact Monoclonal Antibody was analyzed by a standard stock bench-top Orbitrap mass spectrometer