Polymer-based protein engineering has enabled the synthesis of a variety of protein-polymer conjugates that are widely applicable in therapeutic, diagnostic and biotechnological industries. Accurate characterizations of physical-chemical properties, in particular, molar masses, sizes, composition and their dispersities are critical parameters that determine the functionality and conformation of protein-polymer conjugates and are important for creating reproducible manufacturing processes. Most of the current characterization techniques suffer from fundamental limitations and do not provide an accurate understanding of a sample's true nature. In this paper, we demonstrate the advantage of asymmetrical flow field-flow fractionation (AF4) coupled with multiple detectors for the characterization of a library of complex, zwitterionic and neutral protein-polymer conjugates. This method allows for determination of intrinsic physical properties of protein-polymer chimeras from a single, rapid measurement.
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