中文摘要：

目的：探索骨髓间充质干细胞（mesenchymal stem cells，MSCs）三维扩增原位冻存策略实践性及其对传代迁徙和成骨分化能力的影响。方法：制备β—TCP三维支架，相对2D平板考察三维扩增细胞数，冻存后以CCK-8染色计数考察收获冻存效率，迁徙率，并以ALP染色考察总体成骨分化水平和比分化活力。结果：与2D平板相比，3D培养速率提升7倍，原位冻存细胞收获存活效率由传统消化吹打方式的55．9％提升为81.3％；冻存处理后其迁徙率传代效率下降无统计学意义（P〉0．05），比ALP活性经冻存处理非但未下降，反而在迁徙传代支架上显著性提升。结论：利用B—TCP三维支架扩增、传代MSC并进行原位冻存不影响传代迁徙能力，对成骨分化水平反而有贡献作用，具有较好的科研及工业生产应用前景。

英文摘要：

Objective：To explore the practicability of in-situ cryopreservation for mesenchymal stem cells （MSCs） and its effect on cell viability, the later migrating passage and capability of osteogenic differentiation. Methods：β -TCP 3D Scaffolds were produced. MSCs amplification in 3D environment was investigated in comparison with 2D. The harvest and survival rate, as well as the migration rate, were tested by CCK-8 reagent staining for cell number calculation. ALP activity and specific ALP activity were examined for both groups of the scaffold-cells constructs, which had experienced the in-situ cryopreservation or not. Results：The 3D propagation speed was about 7 times than that of the 2D. The harvest and survival rate had risen from 55.9% to 81.3% when the 3D in-situ cryopreservation was applied instead of the traditional 2D trypsinization and cryopreservation. The migration rate of the cryopreservation group was not less than the non-cryopreservation group. Interestingly, the specific ALP activity of the cryopreservation passage group was even higher than the non-cryopreservation group. Conclusions：Employing β -TCP 3D Scaffolds for MSC propagation, passage and in-situ cryopreservation was applicable and promising.