Early and effective integration of a metal implant into bone tissue is of crucial importance for its long-term stability. While different material properties including surface roughness and wettability but also initial blood-implant surface interaction are known to influence this osseointegration, implications of the latter process are still poorly understood. In this study, early interaction between blood and the implant surface and how this affects the mechanism of osseointegration were investigated. For this, blood coagulation on a micro-roughened hydrophobic titanium (Ti) surface (SLA-Hphob) and on a hydrophilic micro-roughened Ti surface with nanostructures (SLActive-HphilNS), as well as the effects of whole human blood pre-incubation of these two surfaces on the differentiation potential of primary human bone cells (HBC) was assessed. Interestingly, pre-incubation with blood resulted in a dense fibrin network over the entire surface on SLActive-HphilNS but only in single patches of fibrin and small isolated fiber complexes on SLA-Hphob. On SLActive-HphilNS, the number of HBCs attaching to the fibrin network was greatly increased and the cells displayed enhanced cell contact to the fibrin network. Notably, HBCs displayed increased expression of the osteogenic marker proteins alkaline phosphatase and collagen-I when cultivated on both surfaces upon blood pre-incubation. Additionally, blood pre-treatment promoted an earlier and enhanced mineralization of HBCs cultivated on SLActive-HphilNS compared to SLA-Hphob. The results presented in this study therefore suggest that blood pre-incubation of implant surfaces mimics a more physiological situation, eventually providing a more predictive in vitro model for the evaluation of novel bone implant surfaces.