Isolation and maintenance of viable sheep hepatocytes

Abstract

Hepatocytes were isolated by collagenase perfusion of the caudate lobe of the sheep liver. The method consistently gave good hepatocyte preparations (viability > 85% by trypan blue exclusion) and the viability of these hepatocytes was maintained for approximately 4 h in media containing defatted bovine serum albumin and gelatin. These proteins were also used during the storage of cells under refrigeration. The isolated hepatocytes were physiologically competent as judged by leakage of the cytosolic enzyme lactate dehydrogenase, by measurement of adenosine-5′-triphosphate concentrations, and by their ability to conjugate phenolphthalein and 4-methyl umbelliferone. These hepatocytes would be valuable in biochemical and toxicological studies relevant to ruminants, such as in research into ‘facial eczema’, a hepatogenous photosensitivity disease which causes the loss of millions of dollars to New Zealand's agricultural economy.