A Controllable Differential Stain for the Hypophysis by Adaptation of the Mallory Triple Stain

A selective and controllable staining method for the hypophysis has been developed with rat material, using Mallory's triple stain as a basis.

Fix in Zenker neutral formol for 6 hours. Longer fixation is undesirable. Transfer to 30% alcohol plus a few drops of a saturated solution of I₂ in aqueous KI over night. Gradually complete dehydration and clear in cedar oil. Infiltrate with a paraffin mixture (paraffin, rubber-paraffin, bayberry wax and beeswax). Section 3-Sμ. Hydrate to distilled water, placing a few drops of a KI-I₂ solution in the 50% alcohol. Stain in 1% acid fuchsia for 30 minutes. Rinse, and differentiate in a weak NH₄OH solution (one drop 28% NH₄OH to 200 cc. HOH). When differentiation is complete, transfer to a 0.5% phosphomolybdic acid solution for 3 minutes, after first stopping the differentiation with a 0.1% HC1 solution and then rinsing with distilled water. Stain for one hour in a solution of: 1% anilin blue, water soluble, 2% orange 6, and 1% phosphomolybdic acid. Rinse in distilled water plus a few cubic centimeters of the stain. Differentiate in 95% alcohol, transfer to absolute alcohol and clear in a mixture of 30% oil of cedar, 40% oil of thyme, 15% absolute alcohol and 15% xylene. Finally, transfer to xylene and mount.