A hybrid coating of biomimetic apatite and osteocalcin

A novel hybrid coating of biomimetic apatite(BAp) and osteocalcin (OC) was prepared by incubating BAp-coated Ti6A14V coupons in an osteocalcin-containing medium. A significant amount (up to 1.0 wt %) of OC was adsorbed by the BAp coating within 3 h of incubation as demonstrated by high-performance liquid chromatography. Characterizations of the hybrid coating with environmental scanning microscopy and X-ray diffraction indicated that protein adsorption does not alter the microstructure of the coating. The presence of OC in the hybrid coating was visualized with fluorescence microscopy using an immuno-labeling procedure. The affinity of OC to the BAp coating was examined using a 20-h elution test in phosphate-buffered saline and only a minimal amount (<10%) of the loaded OC was eluted out. When the coating was fully dissolved in hydrochloric acid solution after elution, about 78% of the loaded OC could be recovered. Enzyme-linked immunosorbent assay and peptide sodium dodecylsulfate-polyacrylamide gel electrophoresis confirmed the integrity and activity of OC molecules throughout the tests. The preliminary cell culture tests showed a significant effect of OC on the attachment and proliferation of osteoblasts. The quick loading profile and high affinity of OC to the BAp coating make it an ideal candidate for the hybrid coating preparation in clinical environment.