Biomolecular charge variant characterization has been a challenge that involves a cumbersome and time-consuming process. This study presents a novel and efficient method for determining binding kinetics and based on the charge heterogeneity of biological molecules. USP mAb 003 was selected as for evaluation owing to its well-characterized charge heterogeneity and suitability as a benchmarking material. First, the charge heterogeneity of USP mAb 003 was analyzed using imaged capillary isoelectric focusing (icIEF) on the CEInfinite systems by Advanced Electrophoresis Solutions, followed by the collection of individual charge variant fractions on the same system. Next, the collected fractions were tested for their ability to bind to FcγRI and FcγRIIIa, as a model to measure how therapeutic antibodies interact with immune cell receptors to evaluate their safety, efficacy, and mechanism of action. To accomplish this, Digital surface plasmon resonance (Digital SPR) was used to measure the binding of each fraction and Fc receptor, requiring only 2 μL of each. Digital SPR offers a key advantage over conventional SPR platforms due to its low sample volume requirements and higher throughput for fractionated samples. Overall, this poster highlights the streamlined fractionation and characterization of charge variants in biotherapeutics. This will help identify critical charge species that impact binding potency to inform the control steps needed in the process development and manufacturing of a biotherapeutic.
