Originally isolated from bone marrow, mesenchymal stromal cells (MSCs) have since been obtained from various fetal and post-natal tissues and are the focus of an increasing number of clinical trials. Because of their tremendous potential for cellular therapy, regenerative medicine and tissue engineering, it is desirable to cryopreserve and bank MSCs to increase their access and availability. A remarkable amount of research and resources have been expended towards optimizing the protocols, freezing media composition, cooling devices and storage containers, as well as developing good manufacturing practices in order to ensure that MSCs retain their therapeutic characteristics following cryopreservation and that they are safe for clinical use. Here, we first present an overview of the identification of MSCs, their tissue sources and the properties that render them suitable as a cellular therapeutic. Next, we discuss the responses of cells during freezing and focus on the traditional and novel approaches used to cryopreserve MSCs. We conclude that viable MSCs from diverse tissues can be recovered after cryopreservation using a variety of freezing protocols, cryoprotectants, storage periods and temperatures. However, alterations in certain functions of MSCs following cryopreservation warrant future investigations on the recovery of cells post-thaw followed by expansion of functional cells in order to achieve their full therapeutic potential.