Membrane ultrastructure modulates immune cell signaling

Membrane lipids modulate the cell membrane ultra‐structure by promoting nanocluster formation, and by influencing the coupling of the membrane to the cytoskeleton. These in turn modulate the cell membrane signaling. Direct observation of the interplay of signaling and lipid nanodomains is challenging due to their small size and dynamic nature (bimFCS), Monte‐Carlo‐Simulations and optical trap based thermal noise imaging (TNI), to characterize the biophysics of the lipid nanodomains Combining total internal fluorescence microscopy with fluorescence correlation spectroscopy into bimFCS, we are able to measure the diffusion of membrane receptors on multiple length scales simultaneously. We characterize the receptor‐nanodomain interaction continuously on single cells while perturbing the cell through receptor dimerization. I will show how bimFCS measurements, combined with defined TNI to confine a single membrane protein to diffuse for seconds in a small area provides sufficient data for high resolutions maps of local diffusion, local attraction potentials and membrane stiffness. Using a GPI‐anchored protein to probe the cell membrane we detect domains of increased membrane stiffness, which also show increased viscosity and are the preferred location for the GPI‐anchored protein.