Guidance for removal of fetal bovine serum from cryopreserved heart valve processing.
Bovine serum is commonly used in cryopreservation of allogeneic heart valves; however, bovine serum carries a risk of product adulteration by contamination with bovine-derived infectious agents. In this study, we compared fresh and cryopreserved porcine valves that were processed by 1 of 4 cryopreservation formulations, 3 of which were serum-free and 1 that utilized bovine serum with 1.4 M dimethylsulfoxide. In the first serum-free group, bovine serum was simply removed from the cryopreservation formulation. The second serum-free formulation had a higher cryoprotectant concentration, i.e. 2 M dimethylsulfoxide, in combination with a serum-free solution. A colloid, dextran 40, was added to the third serum-free group with 2 M dimethylsulfoxide due to theoretical concerns that removal of serum might increase the incidence of tissue cracking. Upon rewarming, the valves were inspected and subjected to a battery of tests. Gross pathology revealed conduit cracking in 1 of 98 frozen heart valves. Viability data for the cryopreserved groups versus the fresh group demonstrated a loss of viability in half of the comparisons (p < 0.05). No significant differences were observed between any of the cryopreserved groups, with or without bovine serum. Neither routine histology, autofluorescence-based multiphoton imaging nor semiquantitative second-harmonic generation microscopy of extracellular matrix components revealed any statistically significant differences. Biomechanics analyses also revealed no significant differences. Our results demonstrate that bovine serum can be safely removed from heart valve processing and that a colloid to prevent cracking was not required. This study provides guidance for the assessment of changes in cryopreservation procedures for tissues.
Cell & Tissue Systems Inc., North Charleston, SC 29406, USA. kbrockbank@celltissuesystems.com
Kelvin G M
KG
Brockbank
Albert E
AE
Heacox
Katja
K
Schenke-Layland
D000818:Animals; D001696:Biomechanical Phenomena; D002417:Cattle; D015925:Cryopreservation / Q000379:methods*; D005333:Fetus / Q000378:metabolism*; D006351:Heart Valves / Q000473:pathology*; D036641:Microscopy, Fluorescence, Multiphoton; D044967:Serum / Q000378:metabolism*; D034421:Sus scrofa; D046509:Tissue Culture Techniques / Q000379:methods*; D014022:Tissue Survival