Automated Enzymatic Deglycosylation of Monoclonal Antibody with the SampleStream Platform.

Engineered monoclonal antibodies have proven themselves as invaluable biotherapeutics used in clinics worldwide. In discovery, development, and production, it is essential that they are accurately validated to ensure their homogeneity, consistency, safety, and effectiveness, which are commonly conducted via mass spectrometric analysis. However, validation processes can be manual, time-consuming, and costly. One hindrance in the analytical workflow is the relatively long time required to deglycosylate native mAbs with PNGase F on the benchtop, which is usually done to reduce the spectral complexity, increase the ionization efficiency, and facilitate the identification of glycosylation sites. To circumvent this obstacle, a workflow on the SampleStream Platform was developed and optimized to automate in-channel PNGase F-mediated deglycosylation of native monoclonal antibody at unprecedented speed, accomplishing 86% deglycosylation in 3 min. The presented workflow offers a promising strategy to reduce discovery and development costs and streamline the characterization of antibody-based biopharmaceuticals.