Abstract: An auto-nucleating device includes a hollow tube containing a crystalline cholesterol matrix therein. The ends of the tube are closed by a membrane that is impermeable to the cholesterol but permeable to liquids contained in a cryopreservation vessel. The auto-nucleating device is disposed within the vessel and provides a site for ice nucleation during freezing of the liquid within the vessel. One such cryopreservation vessel is a flexible vial having a closed port at one adapted to be pierced by a needle to withdraw the liquid within. The opposite end of the vial is initially open to receive the liquid. The end is then sealed to form a closed system for cryopreservation. Another cryopreservation vessel includes an adaptor mounted to a port of a liquid container. The adaptor includes one tubular branch that is closed by a needle septum while another tubular branch includes a barbed fitting for engaging a flexible tube. The flexible term terminates in a needle septum.