The cryopreservation of cells is a fundamental task for a wide range of applications, from basic storage of cells to fertilization. Cells must be protected against damage during freezing by application of specifíc cryoprotectants and freezing/melting protocols. This chapter presents a brief overview of the predominant cryopreservation methods and most common cryoprotectants. Furthermore, it explores the specifics of cryopreservation of stem cells with various types and levels of multipotency. Special attention is dedicated to the effect of cryopreservation on human pluripotent stem cells, including the effect of different cryoprotectants used to prepare fibroblast feeder layer for the cultivation of pluripotent cells. Finally, the chapter concludes with a review of selected methods that have been successfully used to probe the postthaw state of cryopreserved cells through their viability, inner structure, and mechanical properties.