Lipopolysaccharide (LPS), the main component of the cell wall of most gram-negative bacteria, can activate the innate immune response of host cells, and play important roles during the process of recognition, adhesion, metastasis and pathopoiesis of bacteria. Importantly, the structure of LPS determines both the serotype and the pathogenicity of a bacterial infection. Therefore, accurate analysis of its structure is essential for a better understanding of the relationship between LPS structure and its biological effects. Furthermore, reliable determination of its structure might assist in identification of hitherto unknown strains, as well as in the development of novel antibiotic compounds and vaccines. However, the amphiphilic, multi-charged and structural complicated nature of LPS presents a major challenge for its structural analysis. In this review, we summarize recently developed tools for the analysis of bacterial LPS, covering the latest approaches to extracting, separating, purifying and identifying LPS & its oligosaccharide chains.