Validating Intact Mass Analysis Characterization of Therapeutic Antibodies Through Middle-Down Sequencing

CASSS-MS 2023, Chicago, IL, USA
September 5, 2023

While mass spectrometry offers many advantages in biopharmaceutical characterization, methods based on peptide mapping are labor intensive and time-consuming, generating large data sets that require trained and skilled manual interpretation. In addition, the sample preparation process of peptide mapping can create artifactual modifications and deliver results that are not a true reflection of the native state of the molecule. A faster and less complex alternative to characterizing antibodies by peptide mapping is middle-down intact mass analysis, which utilizes novel enzymes that cleave antibodies at a highly specific site above or below the hinge, resulting in well-defined fragments that are amenable to characterization by LC-MS/MS.

We present a fully automated data processing workflow that enables fast and efficient middle-down protein sequencing and simultaneous intact mass analysis of protein biotherapeutics.