ABSTRACT
Objective To investigate the experimental methods of isolation ,culture and identification of osteoblasts from neo-natal New Zealand rabbits in vitro .Methods Two-step enzymatic digestion was adopted to isolate osteoblasts from skull tissue of neonatal New Zealand rabbits to conduct primary cultured .Inverted phase contrast microscope was employed to study the cellular morphology ,acridine orange fluorescent staining was used to detect the cell adhesion function ,methyl thiazolyl tetrazolium(MTT) assay was employed to measure their proliferation ,and Alizarin red and tetracycline staining were used to test their mineralization . Results Primary osteoblasts were successfully obtained .Inverted phase contrast microscopy showed non-adherent cells were round ,while adherent cells were irregular fusiform ,triangular or polygonal .Acridine orange staining showed the nuclei of osteo-blasts green fluorescence ,with good adhesion ability .Good mineralization ability was also demonstrated by tetracycline and alizarin red staining .Osteoblasts possessed good proliferation activity .Conclusion Utilization of two-step enzymatic digestion contributes to getting a lot of osteoblasts with typical morphological features and biological activity in a short time .