The aim of this study was to compare the effectiveness of velvet antler (VA) from different sections for promoting longitudinal bone growth in growing rats. VA was divided into upper (VAU), middle (VAM), and basal sections (VAB). An in vivo study was performed to examine the effect on longitudinal bone growth in adolescent rats. In addition, in vitro osteogenic activities were examined using osteoblastic MG-63 cells. VA promoted longitudinal bone growth and height of the growth plate in adolescent rats. Bone morphogenetic protein-2 (BMP-2) in growth plate of VA group was highly expressed compared with control. The anabolic effect of VA on bone was further supported by in vitro study. VA enhanced the proliferation, differentiation, and mineralization of MG-63 cells. The mRNA expressions of osteogenic genes such as collagen, alkaline phosphatase, and osteocalcin were increased by VA treatment. These effects of in vivo and in vitro study were decreased from upper to basal sections of VA. In conclusion, VA treatment promotes longitudinal bone growth in growing rats through enhanced BMP-2 expression, osteogenic activities, and bone matrix gene expressions. In addition, present study provides evidence for the regional differences in the effectiveness of velvet antler for longitudinal bone growth.