A Simple and Rapid Wholemount Staining Method with Methyl Green-Pyronin G Applied to A Molluscan Brain Preparation

Abstract

Subesophageal ganglia of mollusks [Helix pomatia] were stained as whole mounts with methyl green-pyronin G to display the relative location of individual neurons. Nuclei appear blue, perikarya red. Ganglion cells are exposed by dissection of the connective tissue in snail Ringer, transferred to a fixative of 2.5% glutaraldehyde in 0.1 M Na cacodylate pH 7.1 at 4.degree. C for 12-24 h, and washed in distilled water for 1 1/2 h. They are stained with methyl green-pyronin G for 1/2-1 h, differentiated in 96% ethanol using many rapid changes, transferred to absolute ethanol for 2 1/2 h and cleared in xylene for 3 h before embedding in Depex in a suitable dish. When the Depex has hardened, the preparation can be stored, and is readily available for subsequent examination. The method may be applicable to other invertebrate tissues, and may be useful in preparing objects for teaching purposes.