Chemical Studies on Tumor Tissue: IV. The Staining with Neutral Red of Fresh Preparations of Mouse Tumor Cells

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Abstract
Introduction In studying the chemical properties of individual tumor cells in vitro, it was desirable to know whether the cells under observation were alive or dead; it was also desirable to determine whether the living cells in any given experiment died during the course of observation. Of the various criteria of cell viability—e.g., motility, phagocytosis, fertilization, cell division, vital staining, etc.—which have been employed in the study of various types of single cells, the one selected as most suitable for the purposes of this study was vital staining. The staining with vital dyes of cells of mammalian tissues both in vivo and in vitro has been investigated by many workers. There is general agreement that vital dyes stain the nucleus in dead cells or in severely injured cells. As regards the living cell, however, the findings have not been so uniform (1–5). In the case of neutral red, it has come to be widely accepted that cells treated with this dye often respond as follows. (a) In the living cell the dye is seen in localized areas (granules, vacuoles) in the cytoplasm while the rest of the cytoplasm, as well as the nucleus, appears colorless (6–10). (b) As a cell dies the granules and vacuoles lose their stain, and the nucleus and the cytoplasm become diffusely red (7, 8, 11, 12). (c) In the dead cell the entire nucleus is stained diffusely red and, in addition, the cytoplasm also is usually stained diffusely red (7, 11, 14–16). Although many types of cells react in this fashion with neutral red, other types do not. As various observers (40, 2, 5, 16–28) have noted, cells of one tissue may exhibit numerous stained cytoplasmic bodies, while cells of another tissue may exhibit few stained bodies or none at all. Furthermore, cells of the same tissue under identical conditions may exhibit this variation in response to the dye. These differences in staining behavior are discussed below.

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