Shigella flexneri is an important human pathogen causing shigellosis. Strains of S. flexneri are serologically heterogeneous and, based on O‐antigens, are currently classified into 14 types. Structures of the O‐antigens (O‐polysaccharides) of S. flexneri have been under study since 1960s but some gaps still remained. In this work, using one‐ and two‐dimensional 1H‐ and 13C‐NMR spectroscopy, the O‐polysaccharides of several S. flexneri types were reinvestigated, and their structures were either confirmed (types 2b, 3b, 3c, 5b, X) or amended in respect to the O‐acetylation pattern (types 3a, Y, 6, 6a). As a result, the O‐acetylation sites were defined in all O‐polysaccharides that had not been studied in detail earlier, and the long story of S. flexneri type strain O‐antigen structure elucidation is thus completed. New and published data on the S. flexneri O‐antigen structures are summarized and discussed in view of serological and genetic relationships of the O‐antigens within the Shigella group and between S. flexneri and Escherichia coli.