An individual cell carries out a vast number of processes simultaneously. These range from interacting with the extracellular environment to metabolism to cell division. Physical isolation of these processes allows the cell to carry out activities that require distinct chemical environments or may interfere with each other. Intracellular phospholipid membranes define and enclose the lumen of intracellular organelles and vesicles. They serve as barriers that control the mixing of molecules in one cellular region with another. Specialized functions for each organelle are carried out by lumenal as well as by integral and peripheral membrane molecules. Both lumenal and membrane-associated molecules can be exploited as markers to identify particular types of organelles. The use of markers for intracellular organelles is critical for the interpretation of many experiments in cell biology.