Optical imaging and spectroscopic characterization of subvisible particles in protein therapeutics

The presence of subvisible particles in protein-based pharmaceutics is a critical quality attribute that is highly regulated due to potential risks to product stability, quality, bioavailability, and patient safety. While numerous analytical technologies have been developed to measure and analyze these particles, optical characterization methods are widely used for their simplicity, robustness, and versatility. Selecting the appropriate technique from a vast array of optical spectroscopy and imaging methods can be overwhelming, but it is crucial for successful characterization. For example, compendial methods such as light obscuration are most commonly used but can underestimate particle counts and are unable to provide chemical identification. This review article aims to provide a comprehensive comparison of optical particle characterization techniques, detailing their physical principles, applications, strengths, and weaknesses. We evaluate methods based on elastic light scattering, flow-based imaging, particle tracking, and vibrational spectroscopy. We highlight the inherent trade-off between analytical throughput and information content, aiming to guide the rational selection of analytical tools for the comprehensive characterization of subvisible particles in protein therapeutics.