High-throughput screening is desirable at early stage of formulation development, where material limitation and stringent timeline are common challenges. The objective is to identify 'aggregation hot spots' within a potential formulation space in order to guide fine-tuning work at a later stage. Under a wide range of conditions proteins can undergo different aggregation pathways and kinetics to produce particles with various sizes, structures, morphologies and possible phase transition, and any single assay may not capture all characteristic information1. Therefore a high-throughput methodology requires consideration of the underlying mechanisms to ensure that a proper formulation space is identified for further evaluation. In this study, both thermal melting and constant temperature incubation based on 96-well plate format were employed to extensively screen conditions (pH, ionic strength, temperature, sugar, amino acid, etc.) for multiple monoclonal antibodies. Optical density, light scattering, particle count, and fluorescence spectroscopy were used to determine the onset of aggregation. Fluorescence dyes including Sypro Orange, Nile Red, DCVJ, bis-ANS, and ThT were compared for their sensitivity to detect various aggregates. Statistical analysis of multivariate data allowed examination of possible correlation between apparent melting temperatures and aggregation kinetics, and also provided insight to global patterns1 that exist in antibody aggregation behaviors.1Li, Y et al, Multi-variate approach to global protein aggregation behavior and kinetics: Effects of pH, NaCl, and temperature for alpha-chymotrypsinogen A, 2009, J Pharm Sci. in press

High-throughput screening of monoclonal antibody aggregation behaviors

239th National Meeting (2010)

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