Investigations on nine different mammalian cell lines revealed that permanently growing cells of one morphological class have numerous membrane properties in common which are different or even lacking in the other cell class. With electrophysiological methods it is shown that the sensitivity of the Na+-K+ pump to ouabain is three orders of magnitude higher in the ionically non-coupled epithelioid cells than in the ionically coupled fibroblastoid cells which respond like primary cultures. This is accompanied by considerably higher binding constants for ouabain of the epithelioid cells as was shown by [3 H] ouabain binding and membrane potential measurements. The epithelioid cells also revealed a 50% lower relative amount of phosphatidylethanolamine and a 60-fold less net synthesis of phospatidylinositol. Finally, although primary cultures cannot proliferate without serum, permanent fibroblastoid cells have a reduced serum requirement and permanent epithelioid cells can proliferate without any serum