FETAL BOVINE PRIMARY CULTURE CELLS SUBMITTED TO DIFFERENT NEGATIVE PRESSURES BEFORE STRAW FREEZING

Abstract Cell cryopreservation is an important tool in the preservation of endangered species. Fetal bovine primary culture-obtained cells were subjected to negative pressure (NP) 200, 500 or 800 mbar, just before (NP0h) or 3 hours before (NP3h) freezing into fine (0.25 mL) straws, using 10% DMSO as cryoprotectant. Fresh and frozen fibroblasts non submitted to NP were used as controls. We evaluated cell viability after freezing, cell proliferation curve, and population doubling time (PDT) every 24 hours during 8 days. Data were submitted to Tukey or Chi square test (P≤ 0.05). The average survival of control group (89.8%) and NP500-0h (88.1%) was higher than other groups; the time of PDT was similar in the fresh (27.5 ± 0.35 h), frozen control (30.1 ± 2.3 h) and NP500-0h groups (32.4 ± 1.6 h). The lowest time was observed in the NP800-0h (21.9 h) group. Freezing of bovine fibroblasts into 0.25 mL straws with 10% DMSO enables high survival rates after thawing. The NP modifies the growth curve of cryopreserved cells and the intensities of 200 or 500 mbar, applied just before freezing the cells, allow proliferation curves similar to those obtained with fresh cells.