Analysis of Fatty Acid and Cholesterol Content from Detergent-Resistant and Detergent-Free Membrane Microdomains

The compartmentalization of cellular membranes into discrete membrane microdomains (known as lipid rafts) challenged the original definition of membranes as containing randomly distributed lipid and protein components. The lipid microdomain hypothesis has generated significant controversy and rigorous inquiry based on the attractive idea that such domains concentrate machinery to mediate cellular events such as signaling and endocytosis. As such, numerous studies have used biochemical, cell biological, and biophysical methodologies to define the composition of such domains in a variety of experimental contexts. In this chapter, we describe methodologies to isolate membranes from cell or tissue sources with biophysical/biochemical properties of membrane microdomains that are amenable to subsequent classical or mass spectrometry-based lipid analytical approaches.