[Use of thermitase in the cultivation of embryonal mouse fibroblasts].

Abstract

Thermitase has been investigated as a means for obtaining single cells from tissue material, for enzymatic detachment of cultivated cells from the substrate and rarification of cells with subsequent passaging of mouse embryonic fibroblasts. The action of this enzyme was compared with that of "trypsin for cell cultivation". Tissue digestion showed that thermitase at 1/50 of the enzyme concentration needed with trypsin, was 1.5 fold more effective in yielding single cells. In cell cultivation thermitase is able to detach cells from the substrate at the same low concentration of 0.05 mg enzyme/ml and to give sufficient rarification, without the need of adding complexing agents. Rate of attachment, cell form and multiplication in subcultures corresponded to those after application of trypsin. The best results in cell detachment and rarification and the most uniform cell morphology were obtained with thermitase at a concentration of 0.025 mg enzyme/ml under addition of 4 mM of complexing agent. At that, thermitase proved 50 fold more effective than trypsin. Another advantage of thermitase is its better storage quality at 4 degrees C in dissolved form.