Effects of Ph on Post-Mortem Retention of Trypan Blue Vital Staining in Tissues of Mice

Abstract

Vital staining of aortas from mice injected subcutaneously (daily for 5 days) with trypan blue was studied. In routine paraffin sections elastic membranes were observed to be well stained and other medial elements unstained following fixation in 10% formaldehyde (25% formalin) at pH 7–9. An identical pattern of vital staining was observed in specimens that had been immersed for 48 hr in saline solutions at pH 7–11. Elastic membranes were not stained, but intermembranous connective tissue was stained after the following: (1) fixation in 10% formaldehyde at pH 1–4 and in Lavdowsky's solution (ethanol, formaldehyde, water and glacial acetic acid), pH 2.3–2.8; and (2) immersion in saline for 48 hr at pH 14. Aortic elastic membranes were vitally stained after fixation by intracardiac perfusion with 10% formaldehyde (pH 7–8) but not after perhion with Lavdowsky's fixative (pH 2.3–2.8). Vital staining was limited to medial elastic membranes in sections of fresh aorta made in a cryostat or by a regular freezing microtome. The vital staining (coarse cytoplasmic granules of dye) within macrophages (Kupffer cells and others) and in cytoplasm of renal tubular epithelium was well demonstrated following use of all methods discussed above