Isolation and cultivation of rat liver epithelial cells

Rat liver epithelial cells (rLEC) can be isolated from 8-day old male Sprague-Dawley rats. Briefly, small fragments of neonatal rat livers are incubated for 15 minutes with 4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid (HEPES) buffered trypsin solution [0.25% (v/v)] and washed twice with calcium- and magnesium-free phosphate-buffered saline (PBS) before plating. Elimination of contaminating fibroblasts is accomplished by taking advantage of their faster attachment to plastic culture dishes (plate-and-wait method). Growth medium consisted of Williams’ E medium without glutamine, 10 % (v/v) fetal bovine serum (FBS), 0.68 mM L-glutamine, 50 µg/mL streptomycin sulphate, 7.33 IU/mL benzyl penicillin, 50 µg/mL kanamycin monosulphate and 10 µg/mL sodium ampicillin. Cell cultures are incubated at 37 °C in a 5 % CO2 and 95 % air humidified atmosphere. Growth media is changed completely every 2 days.