During recent years, the role of proteomics in ecology, population biology and evolutionary studies has been growing. It has been used effectively to resolve taxonomic ambiguity in closely related species and for detection of environmental effects on populations. The family Littorinidae (Gastropoda; periwinkles) includes a set of widely distributed species, which are important members of intertidal communities of the seacoasts around the world. Taxonomic relationships among several species, e.g. within the subgenus Neritrema, are still poorly understood. We explored proteomes of three closely related littorinid species (the so called “saxatilis”-group: Littorina arcana, L. compressa and L. saxatilis) from different intertidal levels of one geographic area. The proteome data were obtained using fluorescence-difference 2D electrophoresis (DIGE) followed by mass-spectrometry for protein identification. We showed that L. compressa formed well-supported clade within the "saxatilis" group, while L. arcana and L. sa