New cryoprotective media for cryopreservation of mammal erythrocytes

PURPOSE: To compare the effectiveness of a newly designed multi-component cryoprotective medium (15% PEO-1500, 10% DMSO, 5% 1.2-propanediol, 5% sucrose) with three known single- and two-component ones containing DMSO (20 %), PEO-1500 (30 %), and glycerol-mannitol (30 % and 4 %, respectively) in the cryopreservation of human, equine, bovine and rabbit erythrocytes. METHODS: Cryopreservation was achieved plunging the cryotubes with erythrocyte suspensions into liquid nitrogen (-196°C) at an average cooling rate of 200 deg/min. During the three cryopreservation stages; equilibration, freeze-thaw and final removal of cryoprotectants, the survival of erythrocytes was estimated by the level of hemolysis, measured spectrophotometrically. RESULTS: With the erythrocytes of all investigated mammals the designed medium produced a higher level of survival at all cryopreservation stages. CONCLUSION: The tested multi-component cryopreservation medium, combining both penetrating and non-penetrating cryoprotectants, diminished their concentration and accompanying toxic effects and achieved a higher survival of human and mammal erythrocytes at all stages of their cryopreservation.