Liquid Nitrogen Storage of Hybridoma Cells

Hybridoma and myeloma cell lines can be stored by slowly freezing cells in an appropriate solution of nutrients and a cryoprotectant such as glycerol or dimethyl sulfoxide (DMSO). In this protocol, cells are centrifuged at 4°C, resuspended in cold freezing solution (10% DMSO in FBS), and then transferred to an appropriate freezing vial. The vials are slowly frozen to -70°C in Styrofoam racks and then stored in liquid nitrogen (LN₂). Cells stored in LN₂ will remain viable for years. Once a frozen vial has been removed from LN₂ storage, it should be thawed as described, grown out into log phase, and refrozen.