ROWAN E. MOORE
Biopharmaceutical Chromatography & Mass Spectrometry Center of Excellence, Thermo Fisher
Scientific, 1 Boundary Park, Hemel Hempstead, HP2 7GE. United Kingdom

KELLY M. MCMAHON 
Biopharmaceutical Chromatography & Mass Spectrometry Center of Excellence, Thermo Fisher
Scientific, 1 Boundary Park, Hemel Hempstead, HP2 7GE. United Kingdom

Abstract

Biotherapeutics represent one of the fastest growing sectors of the pharmaceutical industry. The production of recombinant protein pharmaceuticals, such as monoclonal antibodies (mAbs) and antibody-drug conjugates (ADCs) is subject to strict regulatory guidelines. Peptide mapping is an identity test for proteins and is necessary for stability analysis, primary structural characterization and quality assurance (QA). Outlined herein are the most recent advances in protein sample preparation chemistries, ultra-high performance liquid chromatography (UHPLC), mass spectrometry (MS) hardware and intuitive software for the generation of comprehensive, confident peptide maps.

INTRODUCTION

Peptide mapping is a critical workflow in biotherapeutic protein characterization and is essential for elucidating the primary amino acid structure of proteins. It is used for proof of identity, primary structural characterization and quality assurance (QA). 

The International Council for Harmonisation (ICH) Q6B covers the test procedures and acceptance criteria for biologic drug products, and specifies the use of peptide mapping as a critical quality test procedure for drug characterization, used to confirm desired product structure for lot release purposes. 

In order to generate a peptide map, a protein must first be digested into its constituent peptides. Robust separation and identification of the resultant peptides provides insight into a protein’s full sequence; displaying each amino acid component and the surrounding amino acid microenvironment, including disulfide linkage information. Structural characterization at this level also highlights post translational modifications (PTMs) (Figure 1).

Some technologies currently employed for biopharmaceutical peptide mapping are subject to:

•    Irrepro ...