The notion of lipid rafts, while not new, has never been far from controversy (Pike, 2009) , their existence frequently questioned (Munro, 2003) . It is almost 30 years since Karnovsky et al. (1982) suggested that protein diffusion in membranes is not free, but somehow constrained. They went on to detail the now familiar concept of the organization of lipids in domains, which may have functional significance. The differential detergent (Triton X-100) solubility of glycosylphosphatidylinositol (GPI)-anchored proteins and transmembrane proteins was first shown by Hooper and Turner (1988) . Later work showed the importance of lipid rafts in protein sorting and segregation, with GPI anchored proteins being preferentially localized in lipid rafts (Brown and Rose, 1992; Brown and London, 2000; Pike, 2009 ). Other lipid modifications of proteins have also been described, such as palmitoylation and myristoylation which may influence raft localization (Brown and London, 2000; Smotrys and Linder, 2004; Pike, 2009 ). In describing membrane lipid clusters as moving platforms, or rafts, which are enriched in both sphingolipids and cholesterol, perhaps the most important finding was that proteins could be segregated being selectively included or excluded from the rafts (Simons and Ikonen, 1997) . In this way, raft localization can serve to facilitate or obstruct protein interactions (Brown and London, 2000; Lingwood and Simons, 2009) or act as a protein scaffold while allowing diffusion (Maxfield and Tabas, 2005) . Despite significant efforts in developing methodology and techniques for lipid raft research, there are still some shortcomings in precise determination of their size, structure, and composition which were recently critically discussed by Simons et al. (2010) . The most important issue which still induces debate is related to application of detergents for isolation of lipid rafts and methylβ-cyclodextrin for extraction of cholesterol from cell membranes which could lead to formation of artificial complexes not existing in the natural environment. It has now been established that detergent-resistant membranes (DRMs) are not identical to lipid rafts and the data on proteins detected within DRMs from various types of cells should be treated with caution and not always considered to be functionally raft localized (Lichtenberg et al., 2005; Chen et al., 2009a) . Development of a new technique that purifies nano-meso scale DRMs at 37˚C in an ionic buffer that preserves the lamellar phase of the metastable inner leaflet lipids could be a significant step toward purifying individual physiologically relevant rafts (Morris et al., 2011) . Although Triton X-100 is still the standard detergent in preparation of lipid rafts some groups claim that use of Brij 96 produces more reliable results although comparing properties of both detergents in the same set of experiments www.frontiersin.org have not produced significantly different results. Moreover it has been suggested that preparation of the rafts in a buffer mimicking the cytoplasmic environment will preserve the structure of the rafts under physiological conditions by retaining the proteins associated with the intracellular part of the membrane (Chen et al., 2009b) .