Optical Imaging of Protein Aggregation Reactions In Vitro and in Cells

1-Subg Mapping Cell Surface Adhesion by Rotation Tracking and Adhesion Footprinting Isaac T.S. Li1, Yann R. Chemla2, Taekjip Ha3,4. Chemistry, University of British Columbia, Kelowna, BC, Canada, Department of Physics and Centre for Physics of Living Cells, University of Illinois Urbana-Champaign, Urbana, IL, USA, Department of Biophysics and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA, Howard Hughes Medical Institute, Baltimore, MD, USA. Rolling adhesion is the behaviour that leukocytes and circulating tumour cells exhibit as they passively roll along blood vessel walls under flow. It plays a critical role in capturing cells in the blood, guiding them toward inflammation sites, and activating cell signalling pathways to enable their subsequent transmigration. Rolling adhesion is mediated by catch-bond-like interactions between selectins expressed on endothelial cells lining blood vessels and P-selectin glycoprotein ligand-1 found at microvilli tips of leukocytes. Despite our understanding of individual components of this process, how the molecular details of adhesion bonds scale to cell-surface adhesion and rolling behaviour remains poorly understood. Here, we developed 2 label-free methods that map the functional adhesion sites and their strength on a leukocyte surface. The first method relies on tracking the rotational angle of a single rolling cell, which confers advantages over standard methods that track the centre-of-mass alone. Constructing the adhesion map from the instantaneous angular velocity reveals that the adhesion profile along the rolling circumference is inhomogeneous. We corroborated these findings with a second method that allowed us to obtain a footprint of molecular adhesion events using DNA-based molecular force probes. Our results reveal that adhesion at the functional level is not uniformly distributed over the leukocyte surface as previously assumed, but is instead patchy.