Blood vessels are placed in 2 ml Liquid Nitrogen Storage Ampoules (Gibco AG, Basel, Switzerland) filled with 1.6 ml cryomedium (RPMI 1640 culture medium containing 1.8 M dimethyl sulfoxide and 0.1 M sucrose) .
After equilibration for 10 - 20 min at room temperature with the cryomedium the vials are placed into a polystyrene box (11x11x22 cm) and slowly frozen at a mean cooling rate of -0.6 to -1°C/min in a freezer maintained at -70° C. After 2 h the vials are transferred into liquid nitrogen and stored at -196°C until use.
Before use the samples are placed for 10 min on
dry ice and exposed for 6 min to room temperature
before being thawed within 4 min in a 37°C water bath
giving a mean thawing rate of about 15°C/min.
Thereafter the tissues are placed for 2 min in
Krebs-Henseleit solution containing 50% cryomedium
|Cryopreservation of veins|