Dissociation Constant of Integrin-RGD Binding in Live Cells from Automated Micropipette and Label-Free Optical Data

The binding of integrin proteins to peptide sequences such as arginine-glycine-aspartic acid (RGD) is a crucial step in the adhesion process mammalian cells. While these bonds can be examined between purified and their ligands, live-cell assays are better suited gain biologically relevant information. Here we apply computer-controlled micropipette (CCMP) measure dissociation constant (Kd) integrin-RGD-binding. Surface coatings with varying RGD densities were prepared, detachment single cells from surfaces was measured by applying local flow inducing hydrodynamic lifting force on targeted discrete steps. average behavior populations then fit according chemical law mass action. To verify resulting value Kd2d = (4503 ± 1673) 1/µm2, resonant waveguide grating based biosensor used, characterizing fitting kinetics cell populations. Both methods yielded Kd within same range. Furthermore, an analysis subpopulations presented, confirming ability CCMP characterize both whole population levels. introduced methodologies offer convenient automated routes quantify adhesivity living before further processing.