The hydroxyapatite/collagen (HAp/Col) sponge with 95% (v/v) porosity was prepared by freeze-drying of a HAp/Col fiber suspension. MG63 cells were seeded onto the HAp/Col sponge and cultured under a pressure/perfusion condition with osteogenic supplements. A collagen (Col) sponge was used as a control. The cells with sponge were examined by a histology, total DNA content and gene expression. The cells showed good attachment and proliferation everywhere in the HAp/Col sponge, while the cells mainly proliferated at the peripheral part of the Col sponge. Thus, total DNA content in the HAp/Col sponges reached 1.8 times greater than that in the Col sponges at Day 21. Further, the cells and extracellular matrix only in the HAp/Col sponge were calcified, although the cells in both sponge evenly expressed osteogenic gene. These results suggest that the HAp/Col sponge could be useful as a scaffold for bone tissue engineering.