Acidic pH‐Triggered Membrane Fusion Enables Lyophilized Lipid Nanoparticles as Potent Cryoprotectants for Human Erythrocytes

The cryopreservation of human red blood cells (RBCs) is essential for maintaining a stable blood supply, but current glycerol-based methods are hindered by complex and time-consuming post-thaw processing. While trehalose is a promising biocompatible cryoprotectant, its efficient intracellular delivery into non-endocytic RBCs remains a major hurdle. Here, a lyophilized, ionizable lipid nanoparticle (LNP) platform is reported that overcomes this challenge via an acidic pH-triggered membrane fusion mechanism. This approach, inspired by mRNA vaccine technology, enables rapid (<30 min) and efficient intracellular delivery of trehalose. Systematic optimization of the LNP composition and process parameters resulted in a remarkable post-thaw RBC recovery rate of 85%, an efficacy comparable to the clinical glycerol standard but achieved with a simple, single-wash protocol. The efficacy depends on the specific lipid synergy to enable pH-responsive fusion. Critically, the lyophilized LNP powder retains full functionality, ensuring long-term stability and simplifying storage and transportation logistics. This study establishes a clinically relevant, translation-ready cryopreservation strategy that bridges nanotechnology and transfusion medicine, offering a next-generation solution to longstanding challenges in cell preservation.