Modeling the navigating forces behind BSA aggregation in a microfluidic chip

Microfluidic chips are powerful tools for investigating numerous variables including chemical and physical parameters on protein aggregation. This study investigated the aggregation of bovine serum albumin (BSA) in two different systems: a vial-based static system and a microfluidic chip-based dynamic system in which BSA aggregation was induced successfully. BSA aggregation induced in a microfluidic chip on a timescale of seconds enabled a dynamic investigation of the forces driving the aggregation process. This study employed a combination of experimental approaches, including biophysical and microscopic methods, and computational simulations using MATLAB and COMSOL Multiphysics. Obtained results revealed that Brownian movement, advective mixing, and laminar flow applied in favor of the formation of amyloid-like aggregates through the entire pathway. Furthermore, heating provided the necessary energy for the initial BSA's partial unfolding. In the following, space restriction and the cumulative effects of repulsive electrostatic and attractive van der Waals forces contributed to forming BSA clusters as a partially unfolded intermediate in the first few seconds of the aggregation process. Consequently, the synergistic effects of hydrodynamic forces (including shear force), hydrophobic interaction, and space restriction resulted in the deposition of larger aggregates on the channel sidewalls. Due to the elevated local concentration of BSA clusters alongside the strong shear force toward the channel sidewalls, the deposited structures underwent a structural conversion to form amyloid-like aggregates within a few seconds. In this study, we not only elucidated the molecular mechanisms underlying BSA aggregation but also highlighted the forces driving the aggregation process in microfluidic systems, explaining how it occurs within a timescale of seconds.