Cold storage of biopsies from wild endangered native Chilean species in field conditions and subsequent isolation of primary culture cell lines

Abstract

We evaluated the possibility of deriving primary cell cultures from tissue biopsies taken in field conditions from six threaten endemic Chilean species of free-ranging mammals. Biopsies were taken either by ear punching or darts fired to animals and hold in hypothermic conditions (4 degrees C) in defined salt solution for time periods ranging from 0 to 7 d before biopsy samples reached the cell culture laboratory. Previously, holding times were evaluated in experimental cows in controlled conditions. Two enzymatic treatments, collagenase alone or collagenase followed by tripsin, were used to disaggregate tissues for cell culture. We found that ear notches and dart-derived biopsies can be storaged at 4 degrees C for 1 wk and still yield primary cultures. For dart-derived biopsies, there was an invert correlation between length of cold storage and cellular viability in culture. Healthy fibroblast cell lines were obtained in 92% of the biopsies taken despite the origin (punch or biopsy). We are not aware of similar study for free-ranging animals, especially for the use of darting system to biopsy wild terrestrial mammals, we believe that our results could help for a more widespread implementation of these procedures in the practice of ex situ conservation.