[Trypan blue staining capacities of the culture cells (2)--The mechanism of staining].
The cultured cells dispersed by rubber policeman were stained with trypan blue, but these cells were viable at the meaning of keeping viral infective center forming capacity and at the meaning that trypan blue stained cells were transferred to unstained ones by trypsin treatment. This phenomenon may be important not to miscount numbers of the viable cells in cultured cells. Phagocytosis (endocytosis) is well known as a reaction inhibited by NaF or monoiodoacetate, but these metabolic inhibitors did not inhibit trypan blue staining of the cells dispersed both by rubber policeman and 0.05% trypsin treatment. The cells suspended in higher osmotic pressure media were more stainable with trypan blue than lower ones. Addition of K+ to the reaction mixture was not effective on trypan blue staining. These results suggest that "alive" cells dispersed by rubber policeman are stained with trypan blue not to utilize phagocytosis (endocytosis) or Na+-K+ pump, whose reactions couple with energy production. Trypan blue staining capacity of the cultured cells may be depended on pore sizes of protein channels (for example, fixed or gated transport protein channel) in lipid-bilayers of cell membranes. It is concluded that the cells dispersed by trypsin treatment (spherical shape in microscopical observation) and spherical cells in rubber policeman dispersing ones are unstained with trypan blue to contain smaller pore size channels in the cell membranes, but the rest, amoeba-like form cells dispersed by rubber policeman and dead cells are stained to contain larger ones.