Innovations in Platelet Cryopreservation: Evaluation of DMSO-Free Controlled-Rate Freezing and the Role of a Deep Eutectic Solvent as an Additional Cryoprotective Agent

Cryopreservation is a well-established method for extending platelet shelf-life and addressing supply shortages. Traditionally, this involves dimethyl sulfoxide (DMSO) as a cryoprotective agent (CPA), but recent studies suggest that using controlled rate freezing (CRF) with only NaCl may offer a less toxic alternative. To explore further optimization, this study assessed whether adding 10% choline chloride-glycerol, a deep eutectic solvent (DES), could enhance platelet quality in CRF/NaCl cryopreservation. Ten double-dose buffy coat platelet units were divided into test (DES-treated) and control (NaCl-only) groups. After DES exposure (10% for 20 min), all units were prepared using the NaCl protocol and frozen at -80 °C with CRF equipment, then stored for over 90 days. Upon thawing and reconstitution in AB plasma, no significant differences were observed in platelet content post-thaw between control and test units (255 ± 43 vs. 257 ± 41 × 10⁹/unit), post-thaw recovery (>85%): respectively, Δψ (JC-1% pos 63 ± 15 vs. 68 ± 17), LDH (% of total 10 ± 6 vs. 9 ± 6), (CD63% 77 ± 9 vs. 82 ± 7), (CD62P % 72 ± 15 vs. 76 ± 11), (CD42b % 78 ± 9 vs. 80 ± 9), (CD61% 79 ± 9 vs. 78 ± 9), (CD41% 81 ± 11 vs. 83 ± 7), (PAC-1% 33 ± 10 vs. 32 ± 8), (Pecam-1% 78 ± 11 vs. 80 ± 8), (GPIV % 72 ± 10 vs. 74 ± 11), (LAMP-1% 26 ± 14 vs. 11 ± 9), (MPCD61+ % 41 ± 11 vs. 46 ± 10), (ROTEM CT 56 ± 7 vs. 55 ± 6), (ROTEM CFT 110 ± 70 vs. 106 ± 67) and (ROTEM MCF 35 ± 6 vs. 36 ± 6). These findings support the feasibility of CPA-free CRF-based platelet cryopreservation while maintaining functional integrity.