Both approaches are valid and recognised for LC‑MS HCP quantification. The key advantage of spiked‑in proteins is that they are added before digestion and go through the entire workflow, meaning they account for variability in sample preparation and digestion, resulting in more robust and representative quantification.

In contrast, labelled peptides are typically added after digestion and mainly correct for variability in the LC‑MS measurement itself. While they can offer high precision for targeted quantification, they do not capture upstream variability.

In short, spiked‑in proteins are preferred for global, robust HCP quantification, whereas labelled peptides are better suited for targeted, high‑precision analysis of specific proteins. We have written a white paper on this - please feel free to contact barattini@alphalyse.com for a copy.

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.