Galectin-8 (Gal8) interacts with β-galactoside-containing glycoproteins and has recently been implicated to play a role in platelet activation. It has been suggested that Gal8 may also interact with platelet coagulation factor V (FV). This indispensable cofactor is stored in α-granules of platelets via a poorly understood endocytic mechanism that only exists in megakaryocytes (platelet precursor cells). In this study, we now assessed the putative role of Gal8 for FV biology. Surface plasmon resonance analysis and a solid phase binding assay revealed that Gal8 binds FV. The data further show that β-galactosides block the interaction between FV and Gal8. These findings indicate that Gal8 specifically interacts with FV in a carbohydrate-dependent manner. Confocal microscopy studies and flow cytometry analysis demonstrated that megakaryocytic DAMI cells internalize FV. Flow cytometry showed that these cells express Gal8 on their cell surface. Reducing the functional presence of Gal8 on the cells either by an anti-Gal8 antibody or by siRNA technology markedly impaired the endocytic uptake of FV. Compatible with the apparent role of Gal8 for FV uptake, endocytosis of FV was also affected in the presence of β-galactosides. Strikingly, thrombopoietin-differentiated DAMI cells, which represent a more mature megakaryocytic state, not only lose the capacity to express cell-surface bound Gal8 but also lose the ability to internalize FV. Collectively, our data reveal a novel role for the tandem repeat Gal8 in promoting FV endocytosis.