VITAL STAINING OF AMBOCYTE TISSUE OF LIMULUS

1. Neutral red, as a representative of the basic dyes, stains the granules of the ambocytes. The stained granules lose their stain gradually in neutral and almost immediately in acid solutions. Alkaline solutions intensify the staining.

2. In addition to the granules, droplike structures in the ambocytes are stained with neutral red. The latter are very much more resistant to decoloration than the granules. They do not give off spontaneously the stain as readily as do the granules, nor are they as readily decolorized under the influence of acid as are the latter.

3. The effect of acid and alkali on the neutral red stain in ambocytes can be demonstrated not only in single cells microscopically, but also in the test tube if we use ambocyte tissue previously stained with basic or acid stains. Acids cause the giving off of basic dyes, and alkali causes the loss of acid dyes which had previously combined with the tissue. This relation prevails notwithstanding the fact that acid and basic dyes stain different parts of the tissue. Neither heating nor a preliminary treatment of the tissue with acid and alkali alters this result.

4. We conclude from these experiments that vital stains may be taken up by cells in two forms, (1) through the electrostatic forces of primary or secondary valencies they may be attached to the granules or to the cell protoplasm, or, (2) they may be present in a solution in circumscribed areas of the cell. In the latter form they behave towards environmental factors in a way similar to solutions of the stain. It is possible that also the acid vital dyes exist in the latter form, whenever they are taken into a cell.