A Fast and Simple Workflow for Monoclonal Antibody (mAb) Post-Translational Modifications (PTM) Study Using Shimadzu LCMS-9030 Q-TOF

User Benefits

- Simple and straightforward workflow for study on post-translational modifications (PTMs) of mAbs. - High-resolution Q-TOF mass spectrometry LCMS-9030 coupled with Protein Metrics software for data processing can give accurate quantification of amino acid site relative abundance.

Introduction

Antibody therapeutics hold significant growth potential in the biopharmaceutical industry. Currently, there is a growing interest in using liquid chromatography mass spectrometry (LC/MS) to characterize monoclonal antibodies (mAbs) to support early to late-stage drug development. mAbs are macro-proteins that consist of two identical heavy chains and two identical light chains, linked together by disulfide bonds. Each mAb contains a fragment antigen-binding (Fab) region that binds to the antigen and a fragment crystallizable (Fc) region that is at its tail. mAbs can be identified by the unique Complementarity Determining Regions (CDRs) found on the heavy and light chains. In this report, peptide mapping analysis was conducted using NIST monoclonal antibody (NISTmAb) reference material, RM 8671. During the sample preparation, artificial modifications can be introduced to the mAbs such as prolonged exposure to high temperature and incubation conditions. These artificial modifications can increase the oxidation and deamidation relative abundance values. Therefore, it is important to store mAbs correctly and strictly adhere to sample preparation incubation conditions. Trypsin enzyme was used to digest the mAbs into smaller peptide fragments. The digested peptides were then separated using liquid chromatography and their masses were detected using Shimadzu LCMS-9030 QTOF Data- Dependent Acquisition (DDA) mode. Data processing was performed using Protein Metrics software to quantify the amino acid site relative abundance.

December 11, 2023 GMT