Observation of Fluid–Fluid Immiscibility in Ternary Mixtures of DPPC, DOPC and Cholesterol

As discussed chapter 1, cholesterol–rich domains in biomembranes, called rafts, are believed to be involved in many cellular functions [1, 2, 3, 4, 5]. Biochemical studies on the insoluble membrane fraction obtained from detergent extraction of plasma membranes show that it mainly consists of saturated lipids, such as sphingomyelin and cholesterol. The detergent insoluble fraction, known as detergent resistance membranes (DRM), are believed to come from the rafts [6, 7, 8]. Therefore, ternary mixtures of a saturated lipid, an unsaturated lipid and cholesterol have become a popular model system to study the formation and structure of cholesterol–rich domains. We shall refer to these mixtures as ternary raft mixtures. To date, there is no direct evidence for the existence of rafts in plasma membranes. However, in model membranes of raft forming composition, two fluid phases have been found to coexist, one of them rich in the saturated lipid and other rich in the unsaturated one. These two fluid phases are known as the liquid ordered (lo) phase and liquid disordered (ld) phase, respectively, in the literature. Although microscopy and spectroscopy studies show the phase separation, there has been no report of such phase coexistence using diffraction techniques. This discrepancy has been very puzzling and there have been suggestions that the in-plane separation of the two phases may not lead to a macroscopic phase separation [9]. We have carried out x-ray