Dimethyl sulfoxide (DMSO) has long been the gold-standard cryoprotectant for preserving mammalian cells, owing to its superior ability to penetrate cell membranes, suppress intracellular ice formation, and preserve cellular integrity during freeze–thaw cycles. However, growing recognition of DMSO's limitations—including cytotoxicity, induction of molecular and functional alterations, and severe infusion-related adverse events such as neurological and cardiovascular complications—has driven the development of DMSO-free cryopreservation media. These formulations aim to preserve cell viability and functionality while eliminating the risks associated with DMSO, particularly for cell-based therapeutics where DMSO residues may compromise patient safety and require complex post-thaw washing steps.

Applications

DMSO-free cryopreservation media are particularly critical in cell and gene therapy manufacturing, where regulatory requirements demand defined, low-toxicity formulations. The rapid clinical translation of immune cell therapies necessitates cryopreservation strategies that preserve post-thaw viability and functional integrity. Conventional 10% DMSO-based formulations induce cryoinjury by disrupting membrane integrity, altering cytokine and chemokine signaling, and reducing immune effector function.

Tailored cell type-specific cryoprotectant strategies have been optimized: NK cells benefit from low-DMSO, trehalose-supplemented formulations that preserve granzyme B/perforin expression and IFN-γ/TNF-α secretion; CAR-T cells maintain TCR signaling and IL-2/IFN-γ production in serum-free, proline-enriched media; and dendritic cells retain IL-12/IL-10 secretion and co-stimulatory molecule expression in low-DMSO, antioxidant-supplemented formulations.