Effect of Vital Stains on Cultures of Endamoeba Histolytica.

Because of the more or less elaborate fixation and staining methods used to differentiate the types of Amoeba, the clinician has found it impractical to carry this examination out in routine practice. The vital stains suggested themselves as a simple means by which the differentiated points might be accentuated and recognition made easier in the routine examination. References to vital staining of amoeba in the literature are very few. 1

The technique of Sabine was found impractical because of the nature of the culture media or stool specimen. Aqueous solutions of the dyes were prepared from 1/10 to 2%. A capillary drop of the culture and stain were mixed thoroughly on the slide with an applicator. A cover glass is placed firmly over this, with a blotter or cloth to absorb the excess. If the cover glass is not moderately firm, difficulty may be experienced in focusing the oil emulsion. In 3 to 5 minutes the slides may be examined. The following dyes were used: Neutral red; Trypan blue; Janus green; Brilliant cresol blue; and Nile blue; and the effect noted on the motile forms of the amoeba. Staining reactions on the dying or dead organisms were not consistent.

Neutral red appears non toxic up to 1% solution—the motility being very little affected. The dyes were taken up quickly, and give a very clear differentiation between the endoplasm staining pink and the ectoplasm apparently untouched. Recognition under low power in feces as well as cultures, is very easy, the amoeba appearing as pink refractory bodies. Under oil emersion, the nucleus stands out very distinctly, and by careful focusing as the organism flows along, the chromatin granules appear as a thin line about the periphery, the center appearing more or less clear.