Purpose: The purpose of this study was to investigate if particle size and count analysis can be used as a tool for screening formulation options to minimize protein aggregation.
Methods: The model protein used for this study was NIST 8671 mAb lot 14HB-D-002. Test plates were HTD Biosystems iFormualte RS-2 plates. Chemistry variables included pH 5-7.6, ionic strength 0-200 mM NaCl, and 0-10% stabilizer. Particle size and count analysis was performed using the PSS AccuSizer.
Each plate well was reconstituted by adding 160 µL of filtered DI water to each well. Next 40 µL of NIST 8671 protein at 1 mg/mL was added to each well. The plate was heat stress for 24 hours at 60 C. Particle size and count analysis was performed using the AccuSizer and count/mL data was used for multivariate data analysis to correlate formulation conditions to increased or decreased protein aggregation.
Results: Results shown include Pareto analysis to determine if given formulation chemistries were significant, boderline or not significant in changing particle counts. Other results shown include response surface plots of trehalose vs. pH, trehalose vs. NaCl conc., interaction plots of trehalose, pH, NaCl and buffer concentration, and optimization plots showing optimum conditions to minimize aggregation.
Conclusion: Particle count data indicated that the following conditions provided the optimal chemistry to minimize protein aggregation; pH= 6.1-6.3, NaCl conc. = 40-70 mM, trehalose = 7-8%, and buffer = 50 mM. The AccuSizer also provided particle conc/mL > 10 and 25 µm. All samples passed the USP 787 criteria of 6000 particles/container > 10 μm and 600 particles/container > 25 μm