This volume expands upon the previous edition with current, detailed protocols for investigating membranes and their component lipids in artificial membranes, cells, and in silico. Chapters focus on properties of the component lipids, membranes and their biophysical properties, fluorescent probes for studying membranes, sample preparation, physical techniques to study membrane composition, properties , and function, behavior of cholesterol within a bilayer and examination of cholesterol-dependent phase separation. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Methods in Membrane Lipids, Second Edition seeks to aid scientist in further study into membrane lipids.