Abstract
Aggregation is common in protein drug manufacture, and while the effects of protein particulates are under investigation, many techniques applicable for their characterization have been recently developed. Among the methods available to characterize and quantify protein aggregates, none is applicable over the full size range and different methods often give conflicting results. The studies presented here compare two such methods: dynamic light scattering (DLS) and resonant mass measurement (RMM). The performance of each method was first characterized using polystyrene particle size standards (20, 60, 100, 200, 400, and 1,000 nm) over a range of concentrations. Standard particles were measured both singly and in binary mixtures containing 20 nm particles at a fixed concentration (1014 particles/mL) and various concentrations of one of the other particle sizes (i.e., 60, 100, 200, 400, or 1,000 nm). DLS and RMM were then used to detect unknown aggregate content in stressed samples of IgG. Both instruments were shown to have a working range that depends on particle size and concentration. In binary mixtures and polydisperse solutions, DLS was able to resolve two species in a manner dependent on both concentration and particle size. RMM was able to resolve particles above 200 nm (150 nm for protein) at concentrations below 109 particles/mL. In addition, dilution was evaluated as a technique to confirm and quantify the number of particles in solution.
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ACKNOWLEDGMENTS
The authors thank M. Christine Anderson, Dorothy Scott, Christine Stuart, and Nancy Eller, all of CBER, for helpful comments and suggestions. This work was supported by a grant to the National Institute for Pharmaceutical Technology and Education (NIPTE) sponsored by the FDA (“Critical path manufacturing sector research initiative”, U01 FD004275) and also by an appointment (for JK) to the Research Participation Program at the Center for Biologics Evaluation and Research administered by the Oak Ridge Institute for Science and Education through an interagency agreement between the US Department of Energy and the US Food and Drug Administration.
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Panchal, J., Kotarek, J., Marszal, E. et al. Analyzing Subvisible Particles in Protein Drug Products: a Comparison of Dynamic Light Scattering (DLS) and Resonant Mass Measurement (RMM). AAPS J 16, 440–451 (2014). https://doi.org/10.1208/s12248-014-9579-6
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DOI: https://doi.org/10.1208/s12248-014-9579-6