Exploring tricine as a novel red cell cryopreservative: Lessons and future directions

Cryopreservation allows for storage of red blood cells (RBCs) beyond the standard 35-day period. Current glycerol-based methods are labour-intensive and scale-limited in application. Tricine has been identified as a potential alternative cryoprotectant (CPA), demonstrating efficacy in sheep RBC. This study aims to evaluate the biocompatibility and efficacy of tricine in human RBC cryopreservation.

Human and sheep RBCs were exposed to varying concentrations of tricine (2.0-20.0% w/v) or glycerol (20.0-40.0% w/v). Biocompatibility was assessed via 24-h incubation at 4°C, while cryoprotective efficacy was evaluated following freezing in liquid nitrogen, storage at -80°C and thawing at 37°C. RBC recovery was assessed via spectrophotometric estimation of haemolysis.

Tricine was biocompatible, with <1% haemolysis in both species. When frozen, tricine provided significant protection against cryoinjury in sheep RBC, with maximal recovery at 8.0% w/v (42.17% ± 10.96% of RBC recovered). However, tricine lacked cryopreservative efficacy in human RBC, with post-thaw recovery rates on par with those seen following unprotected freezing. Even at the highest performing concentration (10.0% w/v), human RBC recovery remained low (16.08% ± 2.96%), highlighting the ineffectiveness of tricine in preserving human RBC integrity. Further analyses revealed greater hydrophilicity in sheep haemoglobin, which potentially influences freezing tolerance.

Despite promising results within the ovine model, tricine lacks CPA efficacy for human RBC. Species differences in RBC physiology likely contribute to these discrepancies. These findings emphasize the need for rigorous model selection in cryopreservation research and further investigation into CPA mechanisms.