Living cells are characterized by a well-defined and highly-regulated spatiotemporal organization of their biomolecular content. In particular, non-uniformity in molecules concentration (gradients) and distribution (e.g., polarization), is capital in many key biological processes, from apical-basal establishment in epithelial cells to asymmetric division in stem cells, or directional sensing and motility in neuronal growth cones. In bacteria, polarization has also a pivotal role in crucial processes such as cell division, virulence, chemotaxis, motility and adhesion. For most of these phenomena, the spatial organization of membrane proteins and associated lipids, constituting the interface with the surrounding micro-environment, contribute to the specificity of the signaling outcome. Mechanisms driving the spatial organization of the lipid membrane are thus of particular importance to understand biological polarization. For example, microdomains of ordered lipid phase, called "lipid rafts", are hypothesized to recruit specific sets of transmembrane and scaffolding proteins for functional processes. However, the understanding of the underlying mechanisms directing ordered/disordered lipid phase segregation as well as further recruitment of specific lipids and proteins in sub-domains, remains incomplete in particular due to the difficulty of extracting essential parameters in the highly crowded and multi-factorial environment of living cells.