Aggregation and Size Attributes Analysis of Unadsorbed and Adjuvant-adsorbed Antigens using a Multispectral Imaging Flow Cytometer Platform.

Various vaccine quality attributes should be monitored to ensure consistency, potency, purity, and safety of vaccine products prior to lot release. Vaccine particle size and protein antigen aggregation are two important considerations for particle-adsorbed vaccines. In this study, we evaluated the use of imaging flow cytometry as a potential all-in-one platform to measure adjuvant particle size and to detect protein aggregates through a combination of brightfield microscopy, side scatter detection, and fluorescence microscopy. An aluminum phosphate adjuvant was analyzed for size using the brightfield function, and the size measurement was compared against laser diffraction. Heat-induced protein aggregates of either unadsorbed antigens or aluminum phosphate adjuvant-adsorbed antigens were stained with the fluorescent ProteoStat aggregation dye, followed by detection and analysis using a combination of the brightfield and fluorescence microscopy functions. The change in aggregation of unadsorbed antigens was confirmed using dynamic light scattering. These results demonstrate the versatility of the imaging flow cytometry platform for the evaluation of multiple vaccine quality characteristics.