The present in vitro study investigated select functions (specifically, proliferation, synthesis of intracellular proteins, alkaline phosphatase activity, and deposition of calcium-containing mineral) of osteoblasts (the bone-forming cells) cultured on carbon fibers with nanometer dimensions. Carbon fiber compacts were synthesized to possess either nanophase (i.e., dimensions 100 nm or less) or conventional (i.e., dimensions larger than 100 nm) fiber diameters. Osteoblast proliferation increased with decreasing carbon fiber diameters after 3 and 7 days of culture. Moreover, compared to larger-diameter carbon fibers, osteoblasts synthesized more alkaline phosphatase and deposited more extracellular calcium on nanometer-diameter carbon fibers after 7, 14, and 21 days of culture. The results of the present study provided the first evidence of enhanced long-term (in the order of days to weeks) functions of osteoblasts cultured on nanometer-diameter carbon fibers; in this manner, carbon nanofibers clearly represent a unique and promising class of orthopedic/dental implant formulations with improved osseointegrative properties.