Fate of Stressed Oligoclonal Antibodies Tracked by Fluorescence Cross-Correlation Spectroscopy: IgG Aggregation Proceeds across the Species Barrier

Antibody aggregation is a significant challenge in biopharmaceutical development, affecting drug efficacy and safety. This study applies fluorescence cross-correlation spectroscopy (FCCS) to analyze antibody aggregation, offering a novel approach to understanding this critical process. Our results demonstrated that aggregation can occur without species barriers, as evidenced by the correlated diffusion of different types of stressed antibodies. The absence of a species barrier is significant in elucidating aggregation mechanisms and ensuring the quality of therapeutic antibodies, particularly in oligoclonal formulations. Furthermore, we observed that the aggregation propensity of the antibody fragments (Fab and Fc) differed from that of full-length antibodies, emphasizing the need for a comprehensive analysis of both full-length antibodies and their fragments. Additionally, the self-aggregation propensity of fragments does not always align with their heteroaggregation propensity with preformed antibody aggregates. These findings emphasize the importance of advanced analytical methods to track heterogeneous aggregates accurately and provide insights into the aggregation dynamics of therapeutic antibodies under various conditions.