Mass photometry as a robust method for characterizing adeno-associated virus critical quality attributes in gene therapy vector.

The use of adeno-associated viruses (AAVs) as gene therapy vectors is growing, resulting in a pressing need for efficient, reliable AAV analysis methods. Characterizing AAV critical quality attributes (CQAs), such as the abundance of empty or partially filled capsids, is paramount for developing and producing products with high efficacy, low immunogenicity, and low cost. Available methods include analytical ultracentrifugation (AUC)-considered one of the gold standards of AAV analysis technique-and mass photometry-a newer method based on single-particle light scattering. Mass photometry is a fast, low-cost tool that uses little sample and is suitable for environments requiring good manufacturing practices (GMP) compliance. Here, we assess mass photometry's performance in AAV characterization by using it to analyze a sample of empty AAVs, to measure proportions of empty vs. partially filled vs. full AAVs, to quantify loading in samples ranging from 100% empty to nearly 100% loaded, and to measure samples containing transgenes of different lengths. Comparing to AUC data where appropriate, we find that the two methods give comparable results. Based on its accuracy, precision, measurement capabilities, and practical advantages (e.g., speed and low cost), we conclude that mass photometry is an ideal AAV analysis tool.