The effect of variable liposome brightness on quantifying lipid-protein interactions using fluorescence correlation spectroscopy

► A new formalism to measure membrane partition coefficients using FCS is presented. ► Proteins are assumed to distribute among the liposomes according to a Poisson law. ► Presence of vestigial free dye causes an apparent maximal protein bound fraction lower than 100%. ► Lysozyme binding to POPS-containing liposomes is dominantly driven by electrostatic interactions. ► FCS measurements are easily uncoupled from potential interfering protein-mediated liposome aggregation.