Aldehyde fuchsin staining, direct or after oxidation: problems and remedies, with special reference to human pancreatic B cells, pituitaries, and elastic fibers

Abstract

Successful production of aldehyde fuchsin (AF) having the unique properties described by Gomori depends on each of many critical variables. AF made from basic fuchsins which contain mainly rosanilin (C.I. 42510) do not stain properly-fixed pancreatic B cells, pituitary basophils, or elastic fibers in unoxidized sections. AF made from basic fuchsins containing mainly pararosanilin (C.I. 42500) stains these entities strongly. Substances stained by AF without oxidation fall into two classes: 1) nonacidic peptides and proteins, most of which contain half-cystines, and 2) polyanions, particularly when sulfated. Group 2 substances stain rapidly, Group 1 substances stain slowly. Many modifications of aldehyde fuchsin have been described. Modified aldehyde fuchsins (MAFs) differ in the kind of aldehyde and in the amount of aldehyde and hydrochloric acid used in their formulation; they differ also in the temperature and duration of the ripening necessary before they can be used. If microsections are first oxidized by acid permanganate or other oxidant, MAF staining of pancreatic B cells, pituitary basophils and other substances containing cystines is speeded and intensified. Most modified methods prescribe oxidation, but the author's does not. The chemical basis, final result and potential side-reactions of oxidation methods (OXMAF) differ from those of direct methods (DIMAF) such as the author's. DIMAF staining is slower but inherently simpler and less destructive. The time required for optimal staining with DIMAF depends on the potency of the stain, which in turn depends on how the stain was made and its age. Detection of DIMAF--reactive peptides and proteins may be hampered by the strong staining of polyanions. This can be remedied if the polyanions are first stained with Alcian blue (AB) or other durable basic dye of contrasting color resistant to acid ethanol. Experiences with the AB-DIMAF staining of pancreatic B cells, pituitaries and elastic fibers in formalin-fixed human tissues are detailed. Proper control of the variables which affect MAF will insure useful and reliable results either directly or after oxidation. Authors and editors are urged to be more careful hereafter to distinguish the results of DIMAF from those of OXMAF methods. Published reports should always specify the parameters that affect the properties of MAF. In OXMAF methods the steps intervening between oxidation and staining should be spelled out. Such care should help dispel the confusion and uncertainty which cloud the use and reputation of aldehyde fuchsin at present. This unique dye deserves wider and wiser use.