Integrins require the divalent ions magnesium and manganese for ligand recognition. Here we mechanically enforced bond dissociation to explore the influence of these ions on the mechanical strength of the specific bond between α₇β₁ integrin and its pathologically relevant ligand invasin. Upon addition of these cations to the measurement buffer, we observe a pronounced increase in the force necessary to separate integrin and invasin coated beads. Both ions were found to work synergistically. With free invasin in the measurement buffer we furthermore observe that competitive blocking of binding sites overrides the increase in binding strength of individual beads. We show that this is due to a very strong dependence of bond affinity on divalent ions. Our study illustrates the importance of divalent ions for the regulation of force transmission by integrin ligand bonds on the molecular level. Copyright © 2011 John Wiley & Sons, Ltd.

We present an improved analysis of reflection interference contrast microscopy (RICM) images, recorded to investigate model membrane systems that mimic cell adhesion. The model systems were giant unilamellar vesicles (GUV) adhering via specific ligand–receptor interactions to supported lipid bilayers (SLB) or to patterns of receptors. Conventional RICM and dual-wavelength RICM (DW–RICM) were applied to measure absolute optical distances between the biomembranes and planar substrates. We developed algorithms for a straightforward implementation of an automated, time-resolved reconstruction of the membrane conformations from RICM/DW–RICM images, taking into account all the interfaces in the system and blurring of the data due to camera noise. Finally, we demonstrate the validity and usefulness of this new approach by analyzing the topography and fluctuations of a bound membrane in the steady state and its dynamic adaptation to osmotic pressure changes. These measurements clearly show that macroscopic membrane flow through tightly adhered area is possible in our system.