Spider silk has been investigated for decades due to the intriguing mechanical and also biomedical properties of the silk fibers. Previously, it has been shown that recombinant silk proteins can also be processed into other morphologies. Here, we characterized scaffolds made of the recombinant spider silk protein eADF4(C16) concerning their surface interactions with fibroblasts. Studies of BALB/3T3 cells on hydrogels and films made of eADF4(C16) showed low cell adhesion without observable duplication. Electro-spun non-woven scaffolds made of eADF4(C16), however, enabled both their adhesion and proliferation. Since eADF4(C16) lacks specific motifs for cell attachment, fibroblasts cannot generate focal adhesions with the material's surface, and, therefore, other cell–interface interactions such as topographical anchorage or cell attachment mediated by adhesion of extracellular matrix proteins are discussed in this paper. On non-woven meshes protrusion of filopodia and/or lamellipodia between individual fibers increase the surface contact area, which depends on the diameter of the fibers of the non-woven meshes. In contrast, at flat (film) or microstructured surfaces (hydrogels) such interactions seem to be precluded.