Initiation of Primary Cell Cultures from Lake Trout (Salvelinus namaycush) by a Collagenase Perfusion Technique
IAAAM 1994
Kandal Harr1; James Jacobs2; Paul R. Bowser1
1Department of Avian and Aquatic Animal Medicine, 2Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY

A technique to isolate hepatocytes for primary cell culture was developed to study the etiology of epizootic epitheliotropic disease (EED) in lake trout (Salvelinus namaycush). Herpes-like viral particles associated with this disease have been visualized in extracts of epithelium, kidney, and liver of affected fish. In order to isolate and characterize the virus and develop diagnostic techniques to identify the virus in the field, it is necessary to grow infectious virus in cell culture. The technique used for the isolation of hepatocytes was based on methods developed for mammalian species involving the introduction of dilute collagenase through a venous canula. Hepatocytes were obtained in large numbers with 99.8% viability in pure culture. Hepatocytes were grown on a variety of adhesion surfaces including fibronectin, laminin I and IV, and matrigel at different concentrations under different environmental regimes. Growth was best on fibronectin and 15% matrigel at 17°C. The applications of these techniques for understanding this disease and other possible systems will be discussed.

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Kendal E. Harr

MAIN : Session V : Primary Cell Cultures
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