Typically, two or more well‑behaved peptides per protein are recommended, as this allows cross‑verification and reduces the risk of bias from peptide‑specific effects such as variable digestion or ionisation. Where possible, using additional peptides can further increase confidence.

For global HCP workflows, however, the focus is less on a fixed number per protein and more on leveraging many peptides across the dataset, which helps average out variability and supports robust quantification.

Thomas Kofoed, PhD, CEO of Alphalyse in Denmark, discussed key factors in implementing GMP-compatible LC-MS methods for release assays and highlighted successful cases where this approach has replaced traditional ELISA methods. Don’t miss his presentation and expert insights at the panel discussion.