The ability of natural killer (NK) cells to mediate potent antitumor immunity in clinical adoptive transfer settings relies, in large part, on their ability to retain cytotoxic function following cryopreservation. To avoid potential systemic toxicities associated with infusions of NK cells into patients in the presence of dimethylsulfoxide (DMSO), interest in alternative cryoprotective agents (CPAs) with improved safety profiles has grown. Despite the development of various sugars, amino acids, polyols, and polyampholytes as cryoprotectants, their ability to promote protection from intracellular cryodamage is limited because they mostly act outside of the cell. Though ways to shuttle cryoprotectants intracellularly exist, NK cells' high aversity to manipulation and freezing has meant they are highly understudied as targets for the development of new cryopreservation approaches. Here, the first example of a safe and efficient platform for the intracellular delivery of non‐DMSO CPAs to NK cells is presented. Biocompatible chitosan‐based nanoparticles are engineered to mediate the efficient DMSO‐free cryopreservation of NK cells. NK cells cryopreserved in this way retain potent cytotoxic, degranulation, and cytokine production functions against tumor targets. This not only represents the first example of delivering nanoparticles to NK cells, but illustrates the clinical potential in manufacturing safer allogeneic adoptive immunotherapies “off the shelf.”