THE MECHANISM OF VITAL STAINING WITH BASIC DYES. BY MORTON McCUTCHEON

That intravitam staining with basic dyes occurs more readily in alkaline than in acid solutions has long been known. Among the explanations which have been offered to account for this fact two deserve especial mention. The first is that originally proposed byOverton (1), accepted by Harvey (2), and supported in certain respects by the experiments of Robertson (3) (who, however, on other grounds arrived at a theory diametrically opposed to that of Overton.) According to Overton's theory, cells are freely permeable to the lipoid-soluble dye base but not to the relatively lipoid-insoluble salts of this base with various acids. In acid solutions the dye is present chiefly in the form of salts which enter cells with difficulty, while in alkaline solutions it exists largely in the form of the readily penetrating free base. The difference in staining in the two cases, according to this theory, is due primarily to factors outside of the cell. The second view, advocated in recent years among others by Bathe (4), Rohde (5), and Irwin (6, 7), depends on the fact that in relatively alkaline solutions proteins combine readily with basic and not with acid dyes and show the reverse behavior in more acid solutions. (In this connection, see also Loeb, (8).) According to this view the state of the dye in th~ external solution is of less importance than the condition of the proteins within the cell. If it were possible to vary experimentally and independently of one another the pH inside and outside of a cell it ought to be possible to decide which of these two theories is more in accord with the facts; since if the first i s correct, the degree of staining ought to depend largely on the pH of the external solution, while if the second is correct it ought to depend more on that of the cell itself. More specifically, if two cells, one relatively more alkaline internally than the other, 501