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A dual enzyme method for the establishment of long‐ and medium‐term primary cultures of epithelial and fibroblastic cells from Atlantic salmon gills
Author(s) -
Butler R.,
Nowak B. F.
Publication year - 2004
Publication title -
journal of fish biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.672
H-Index - 115
eISSN - 1095-8649
pISSN - 0022-1112
DOI - 10.1111/j.0022-1112.2004.00521.x
Subject(s) - biology , fibroblast , salmo , collagenase , fibronectin , epithelium , trypsinization , cell culture , matrigel , microbiology and biotechnology , monoclonal antibody , trypsin , staining , gill , cell , enzyme , immunology , antibody , biochemistry , fishery , fish <actinopterygii> , genetics
A dual enzyme disaggregation method using collagenase and then trypsin was developed that allowed the reproducible initiation of primary cultures from Atlantic salmon Salmo salar gills. Cultures had both epithelial and fibroblast morphology and persisted for an average of 20 passages. Growth was dependent upon a minimum concentration of 5% foetal calf serum (FCS) for fibroblasts and 10% FCS for epithelial cells. Growth was mostly independent of substrate, although epithelial cells showed increased growth on type I collagen gels. Matrigel™ cell culture substrate produced reduced growth of fibroblasts and did not benefit epithelial cell growth. Epithelial cells reacted with monoclonal antibodies (MAbs) against mammalian cytokeratins, and fibroblast cells reacted with MAbs against mammalian fibronectin and type I collagen. The method also produced two long‐term cultures: one epithelial and one fibroblast that have been designated RGE‐2 and RGF respectively.