Surface-induced blood clot and thrombus formation following a cascade of coagulation reactions are one of the biggest challenges of those blood-contacting biomaterials (BCB) [1]. Also, the high rate of device-related infections due to the pathogenic adhesion and subsequent biofilm formation is interrupting the successful usage of BCB [2]. Titanium (Ti) and its alloys are widely used as biomaterials and components, especially in cardiovascular applications, due to their good fatigue strength, machinability, formability, and corrosion resistance [3]. Heparin is extensively used as an anticoagulant drug in the clinic, but it also has the ability of cell-growth stimulation and plasma clearing ability [4]. Within this study, alternating current electrophoretic deposition (AC-EPD) has been used for the first time to biofunctionalize Ti substrates surfaces with heparin. A comparison was made with the simple dipping method. Heparin-modified Ti substrates either processed by AC-EPD (Ti-Hep) or simple dipping (Ti-Hepd) were investigated in terms of their resistance against non-specific protein adhesion, anti-bacterial properties against both gram-positive (S. aureus, S. epidermidis) and gram-negative (E. coli) bacteria strains, hemocompatibility (hemolysis and platelet adhesion) and cytocompatibility studies with fibroblast cells (cytotoxicity and cell adhesion), respectively. It was found that for Ti-Hep substrates, non-specific protein adhesion decreased almost fourfold after modification with heparin, while the hemolysis ratio also decreased more than twofold and platelet adhesion was lowered. On the other hand, Ti-Hep surfaces were 25-35% more effective against both gram-positive and gram-negative bacterial biofilm formation compared to bare Ti surfaces, while these did not induce cytotoxic effects on fibroblast cells.