An improved method for the isolation of hepatocytes from liver slices: Selective removal of trypan blue-dyeable cells
✍ Scribed by Peter Bellemann; Rolf Gebhardt; Dieter Mecke
- Publisher
- Elsevier Science
- Year
- 1977
- Tongue
- English
- Weight
- 512 KB
- Volume
- 81
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
An improved nonperfusion method for the isolation of hepatic parenchymal cells using a modified technique of collagenaseihyaluronidase digestion with liver slices is described. The yield of viable cells was considerably increased (30 x IO6 cells/g of liver) by bubbling all solutions with carbogen (95% 0, and 5% CO,) and by adding 100 mg% of glucose to all buffers used. Trypan blue-dyeable cells were selectively removed by a short trypsin-DNase incubation following the isolation procedure, and, thereby, the viability index was conspicuously high (>96'%). The hepatocytes showed normal ultrastructure, and the cell suspension preserved their hormone sensitivity, as tested by an insulin stimulation of the oxidative glucose metabolism. This simple method for isolating large numbers of viable cells may potentially be important for use with small liver pieces. such as biopsy samples. and it is suitable for maintaining hepatocytes in primary monolayer cultures in order to study enzyme regulation at the cellular level in vitro.