Antibody biotherapeutic measurement from pharmacokinetic studies has not been traditionally based on intact molecular mass as is the case for small molecules. However, recent advancements in protein capture and mass spectrometer technology have enabled intact mass detection and quantitation for dosed biotherapeutics. A bioanalytical method validation is part of the regulatory requirement for sample analysis to determine drug concentration from in-life study samples. Results/methodology: Here, an intact protein LC–MS assay is subjected to mock bioanalytical method validation, and unknown samples are compared between intact protein LC–MS and established bioanalytical assay formats: Ligand-binding assay and peptide LC–MS/MS. Discussion/conclusion: Results are presented from the intact and traditional bioanalytical method evaluations, where the in-life sample concentrations were comparable across method types with associated data analyses presented. Furthermore, for intact protein LC–MS, modification monitoring and evaluation of data processing parameters is demonstrated.

The COVID-19 pandemic has resulted in the development of mass spectrometry based methods to characterize, identify, and quantify proteins. These methods are aimed at understanding the structural biology and interaction mechanisms of SARS-CoV-2, or as a complementary method to detect relevant markers. Targeted mass spectrometry, through the detection of viral peptides in proteolytically digested body fluids, has been suggested as a SARS-CoV-2 detection method. The work presented here demonstrates application of the MassLynx Skyline Interface for automated peptide Multiple Reaction Monitoring selection and optimization with a Xevo TQ-XS Tandem Quadrupole Mass Spectrometer.