The behavior of MC3T3 osteoblasts and human bone marrow stem cells on non-coated and hydroxyapatite (HAP)-coated carboxymethyl chitosan (CMCS) scaffolds was investigated in this study. Four HAP-coated scaffolds with different coating morphology and coverage were prepared by mineralization for 1 week in four different mineralizing solutions. Viability, attachment, proliferation, and differentiation of the osteoblasts on these scaffolds were evaluated, and an osteogenic gene expression analysis was carried out to investigate the osteoblastic differentiation of the stem cells. No cytotoxic effects were observed with both the non-coated and coated scaffolds. The non-coated CMCS scaffold supports attachment, proliferation, and differentiation of the osteoblasts and directs stem cell differentiation to osteoblast. Coating the scaffold with HAP substantially enhances these effects on the osteoblasts and stem cells. The main improvement was in the late stage of osteoblast differentiation since osteoblastic differentiation of the osteoblasts and stem cells in this stage was significantly enhanced by the coatings regardless of the variation in morphology and coverage. On the other hand, high HAP coverage was beneficial in stimulating osteoblast attachment and proliferation. This study demonstrates the good potential of HAP-coated CMCS scaffolds as osteogenic scaffolds to stimulate bone healing. (C) 2011 Elsevier Inc. All rights reserved.