Lipopolysaccharides are the major components on the surface of most Gram-negative bacteria, and recognized by immune cells as a pathogen-associated molecule. They can cause severe diseases like sepsis and therefore known as endotoxins. Lipopolysaccharide consists of lipid A, core oligosaccharide and O-antigen repeats. Lipid A is responsible for the major bioactivity of endotoxin. Because of their specific structure and amphipathic property, purification and analysis of lipopolysaccharides are difficult. In this chapter, we summarize the available approaches for extraction, purification and analysis of lipopolysaccharides.