Calibration Standards. Concentration calibration standards were made from lipid mixtures dissolved in chloroform containing a known mol % of labeled lipid. Calibration standards used to quantify labeled lipid concentrations in ternary SLBs contained 20 mol % cholesterol, as this is present in the ternary mixture. Calibration curves without cholesterol were also prepared so that labeled lipids concentrations in mixtures without cholesterol could quantified. The prepared lipid mixtures dissolved in chloroform were spread on plasma cleaned NanoSIMS substrates. The chloroform was then allowed to evaporate to form a lipid film. Concentration calibration standards were kept in a desiccator until use.