Characterization of therapeutic proteins by membrane and in-gel tryptic digestion

Smales, C. Mark and James, David C., eds. (2005) Characterization of therapeutic proteins by membrane and in-gel tryptic digestion. Methods in Molecular Biology, 308 . Humana Press, 482 pp. ISBN 9781588293909. (The full text of this publication is not available from this repository)

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Abstract

The fate and effectiveness of a therapeutic protein product within a patient is, in part at least, determined by potential product variation that leads to differential activity and the half-life of the protein reagents in vivo, and these variations can also contribute towards antigenicity. To assure therapeutic product quality and authenticity, it is therefore necessary to have in place a (or several) method(s) to characterize the possible variation in therapeutic protein products. This is required to determine whether variation in the detailed molecular composition of complex biopharmaceuticals is controlled during bioprocessing. One traditional approach to therapeutic protein characterization involves the proteolytic digestion of the protein of interest in solution followed by high-performance liquid chromatography (HPLC) analysis of the resulting peptide mixture on a reverse-phase column (e.g., see refs. 1,2). The peptides are then usually collected either manually or by a fraction collector, concentrated, and analyzed by either mass spectrometry or N-terminal Edman sequencing. However, such an approach requires relatively large amounts of protein (up to milligrams) and is time-consuming.

Item Type: Edited book
Additional information: Journal Article United States
Uncontrolled keywords: Chromatography, High Pressure Liquid *Electrophoresis, Gel, Two-Dimensional Gels Mass Spectrometry Peptide Mapping/*instrumentation/*methods Polyvinyls Proteins/chemistry/*metabolism/*therapeutic use Trypsin/*metabolism
Subjects: Q Science
Divisions: Faculties > Science Technology and Medical Studies > School of Biosciences
Depositing User: Mark Smales
Date Deposited: 04 Sep 2008 23:15
Last Modified: 30 Apr 2014 11:07
Resource URI: http://kar.kent.ac.uk/id/eprint/6221 (The current URI for this page, for reference purposes)
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