Fig. 4: Multiple lipid probes partition away from protein-rich regions.

a Representative microscopic images showing regions with three different brightness from protein and lipid channels. 1 µM of his-RGG labeled with Atto 488 was applied to both sides of the membrane. Membrane composition: 85 mol% DOPC, 15 mol% DGS-Ni-NTA, and 0.5 mol% Texas Red-DHPE. Scale bar, 5 µm. b Relative intensity profile along the dotted line (from A to B) in the merged channel in a, where green and red lines indicate relative intensity from the protein and lipid channels, respectively. See the caption of Fig. 3c for the definition of relative intensity. c Representative microscopic images after the addition of 1 µM of his-RGG to the top side only of each membrane containing either head labeled (Texas Red-DHPE, Oregon Green-DHPE, and NBD-DHPE) or tail labeled (BODIPY TR-Ceramide and NBD-PC) lipid probes. His-RGG was labeled with Atto 488 or Atto 594, depending on the lipid probes used. The relative fluorescence intensity profile along the dotted line in each merged channel image is shown at the bottom, where gray and black lines represent the intensity from protein and lipid channels, respectively. Buffer: 25 mM HEPES, 100 mM NaCl, pH 7.4. Membrane composition: 85 mol% DOPC, 15 mol% DGS-Ni-NTA, and 0.5–1.0 mol% lipid probe. Scale bars, 50 µm. Source data are provided as a Source Data file.