To readily supply seed cells for tissue engineering and ensure their constant availability for experiments, it is imperative to establish an in-situ cryopreservation method for cell storage. We investigated the effects of a ${eta}$-tricalcium phosphate (${eta}$-TCP) 3D scaffold in-situ cryopreservation method on the migration rate and osteogenic ability of mesenchymal stem cells (MSCs). Compared to using a 2D plate culture and trypsinized cryopreservation, MSCs on ${eta}$-TCP 3D scaffolds demonstrated a higher amplification rate, and the harvest and survival rates (HSR) increased from 55.9 to 81.3% when the 3D in-situ cryopreservation method was applied. The cell migration rate and alkaline phosphatase (ALP) activity were unaffected after in-situ cryopreservation, and unexpectedly, the Specific ALP activity of migrating cells was higher than that of non-cryopreserved cells, suggesting that the cell-scaffold combination could be cryopreserved using the present protocol without loss of proliferative or osteogenic potential. These findings highlight a methodology for 3D scaffold in-situ cryopreservation and passage for MSC production in bone tissue engineering, and present the possibility of designing a perfusion cells/scaffold factory for scale-up production.