Determining the Stoichiometry of a Protein-Polymer Conjugate Using Multisignal Sedimentation Velocity Analytical Ultracentrifugation.

Advances in polymer science have broadened the applications of protein-polymer conjugates as biopharmaceuticals and biotechnology reagents. The complex nature of these conjugates makes characterization challenging. Here, we describe the use of multisignal sedimentation velocity analytical ultracentrifugation to measure the polymer-to-protein ratio. To demonstrate the principle, we applied this approach to a series of antibody-drug conjugates with different polymer-to-protein ratios and various degrees of heterogeneity, and validated results with orthogonal analytical techniques. We found that multisignal sedimentation velocity can provide accurate information on key attributes including polymer-to-protein ratio, which is important for maximizing the therapeutic potential of future protein-polymer conjugates.