Sulfatides are sphingolipids commonly found at the surface of most of eukaryotic cells. Sulfatides are not just structural components of the plasma membrane but also participate in a wide range of cellular processes including protein trafficking, cell adhesion and aggregation, axon-myelin interactions, neural plasticity, and immune responses, among others. The intriguing question is how can sulfatides trigger such cellular processes? Their dynamic presence and specific localization at plasma membrane sites may explain their multitasking role. Crystal and NMR structural studies have provided the basis for understanding the mechanism of binding by sulfatide-interacting proteins. These proteins generally exhibit a hydrophobic cavity that is responsible for the interaction with the sulfatide acyl chain, whereas the hydrophilic, negatively charged moiety can be found either buried in the hydrophobic cavity of the protein or exposed for additional intermolecular associations. Since sulfatides vary in their acyl chain composition, which are tissue-dependent, more emphasis on understanding acyl chain specificity by sulfatide-binding proteins is warranted. Importantly, changes in cellular sulfatide levels as well as circulating sulfatides in serum directly impact cardiovascular and cancer disease development and progress. Therefore, sulfatides might prove useful as novel biomarkers. The scope of this review is to overview cell functions and mechanisms of sulfatide recognition to better understand the role of these lipids in health and disease.