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DIAZO-SAFRANIN FOR STAINING ENTEROCHROMAFFIN

R. D. LILLIE 1, H. J. BURTNER 1, and J. P. GRECO HENSON 1

1 Laboratory of Pathology and Pharmacology, National Institute for Arthritis and Metabolic Diseases, National Institutes of Health, Bethesda 14, Maryland

Freshly diazotized safranin O colors enterochromaffin cell granules black. Background staining can be kept down to a fairly light red. Fairly stable acid aqueous safranin solution and normal sodium nitrite solution can be kept on hand and mixed ex tempore to prepare the diazo solution. A diazotization time of 15 minutes at 3°C is adequate, and the coupling (staining) time of 5 minutes should not he exceeded. Both the specific and the background stains are acid fast. A 1:40 dilution in M/10 Na2HPO4 produces the desired grade of alkalinity and a proper dilution for best. contrast.

In vitro alkaline coupling reactions with diazo-safranin produced colored precipitates with phenol, cresols, naphthols, resorcinol, hydroquinone, pyrogallol and tyrosine. Definite colors were produced on gelatin paper impregnated with resorcinol and the naphthols, less definite with phenol and the cresols. Gelatin paper models gave positive reactions with diazotized agr-naphthylamine for catechol, pyrogallol and phloroglucinol as well as for resorcinol and the naphthols.

The positive coupling with diazo-safranin speaks against a catechol or hydroquinone structure and supports the thesis of a resorcinol structure.

Submitted on January 27, 1953


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