Advancing Intra-Diffusion of Peptide Colloids in Cell-Clustered Spheroids for Enhanced Cryo-Recovery

3D cellular models have been increasingly integrated into the drug development process, but maintaining their functional integrity over extended periods remains a major barrier to clinical translation and commercialization. Cryopreservation represents a key strategy for enabling long-term storage of intact 3D cellular systems; however, the limited diffusion of conventional agents into densely organized structures often compromises post-thaw recovery. We introduce a colloidal solution containing nanoscale peptide-colloid hybrids designed to preferentially interact with the extracellular matrix. This solution infiltrated spheroids and distributed homogenously through 3D spheroidal space, which outperformed conventional cryoprotectants in preserving post-thaw structural integrity and functional activity. At sub-picomolar colloidal concentration, the solution leverages a synergetic Gibbs effect to generate an ice-melting thermodynamic efficiency comparable to high-concentration colligative dimethyl sulfoxide (DMSO). Remarkably, the colloid nanoparticles can be readily removed from the spheroids with a single step of post-thaw rinsing, mitigating the risks associated with residual cytotoxicity.