Abstract：Deer antlers are the only known mammalian organs which can periodically regenerate from pedicles. Antler regeneration is known as a stem cell-based process and antler stem cells reside in pedicle periosteum. In this study conditions of two-dimensional electrophoresis for antler stem cells in sika deer (Cervus nippon) were optimized in the following four aspects including cell processing, staining, isoelectric focusing condition and protein purification. Results showed that comparing with ultrasonic, Bullet Blender had a better effect on cell breaking. It could get more and clearer protein points by combining coomassie brilliant blue staining and silver staining. When the total volt-hours of isoelectric focusing was at 15000 volt-hours, the vertical stripes were relatively lighter. By comparing six kinds of protein extraction and purification methods, the one combining the manual lysate and 2D cleanup kit would get more protein points, lighter background and clearer map. After the comprehensive optimization of two-dimensional electrophoresis, the acquired maps had more and clearer protein spots, less stripes and better reproducibility, meanwhile, they could accord with the requirements of software analysis and experimental treatments. The optimization could apply to the study of antler stem cell proteome. All these researches could provide foundation for the study of comparative proteomics in different developmental phases of antler stem cells.
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