“Lab of the Future”─Today: Fully Automated System for High-Throughput Mass Spectrometry Analysis of Biotherapeutics

Abstract

Here we describe a state-of-the-art, integrated, multi-instrument automated system designed to execute methods involved in mass spectrometry characterization of biotherapeutics. The system includes liquid and microplate handling robotics and utilities, integrated LC-MS, along with data analysis software, to perform sample purification, preparation, and analysis as a seamless integrated unit. The automated process begins with tip-based purification of target proteins from expression cell-line supernatants, which is initiated once the samples are loaded onto the automated system and the metadata are retrieved from our corporate data aggregation system. Subsequently, the purified protein samples are prepared for MS, including deglycosylation and reduction steps for intact and reduced mass analysis, and proteolytic digestions, desalting, and buffer exchange via centrifugation for peptide map analysis. The prepared samples are then loaded into the LC-MS instrumentation for data acquisition. The acquired raw data are initially stored on a local area network storage system that is monitored by watcher scripts that then upload the raw MS data to a network of cloud-based servers. The raw MS data are processed with the appropriately configured analysis workflows such as database search for peptide mapping or charge deconvolution for undigested proteins. The results are verified and formatted for expert curation directly in the cloud. Finally, the curated results are appended to sample metadata in the corporate data aggregation system to accompany the biotherapeutic cell lines in subsequent processes.

Facilitating high throughput bispecific antibody production and potential applications within biopharmaceutical discovery workflows

“…Preparing large bsAb panels through combinatorial approaches such as cFAE typically requires the use of robotic liquid and microplate handling to ensure accuracy and minimize hands-on sample preparation time. Waldenmaier et al 150 have recently reported a fully automated mAb characterization workflow to support cell line selection, starting with mAb purification from cell-line supernatant, prior to intact, reduced, and peptide mapping LCMS analyses and data processing. The sample preparation stages of this procedure could potentially be modified to analyze panels of > 1000 bsAbs to support early screening campaigns, providing both time savings and improved data quality and consistency.…”

An Online Two-Dimensional Approach to Characterizing the Charge-Based Heterogeneity of Recombinant Monoclonal Antibodies Using a 2D-CEX–AEX–MS Workflow

“…These modifications can affect the charge of the protein, resulting in different forms of mAbs with varying charge properties. Therefore, it is essential to monitor charge heterogeneity during the production process to ensure that the final product is of the required quality. , …”