[Histonet] Pinkus' acid orcein Giemsa stain

Bob Richmond rsrichmond <@t> gmail.com
Sat Jun 2 13:10:49 CDT 2012

Luis Chiriboga (at NYU) asks for a method for Pinkus' acid orcein Giemsa stain.

I haven't performed this stain or seen it done, but I copied the
method out of Pinkus' book quite a few years ago. I don't know if
anybody is still doing this stain. Orcein is a natural dye (from a
species of lichen) which can be synthesized, though the synthetic dye
is different from the natural one. Pinkus specified the synthetic and
gave a source (the old Harleco).

Bob Richmond
Samurai Pathologist
Knoxville TN
Mehregan, Amir H. (Wayne State). Pinkus’ Guide to
Dermatohistopathology. 4th ed. Appleton-Century-Crofts, 1986.
Hermann Pinkus, 1905-1985

Acid orcein and Giemsa stain: from
Pinkus H, Hunter R. Simplified acid orcein and Giemsa technique for
routine staining of skin sections. Arch Dermatol 82:699, 1960
Krobock E, Rahbari H, Mehregan AH. Acid orcein and Giemsa stain.
Modification of a valuable stain for dermatologic specimens. J Cutan
Pathol 5:37, 1978.

Fix in formalin or alcohol, without chromium or mercury.

1. Stain in ORCEIN for 30 minutes. Synthetic orcein stains elastic
fibers specifically, with very little background staining. The
background may be decolorized by short immersion in absolute alcohol
or 0.1% acid alcohol.
2. Wash in running water 10 minutes.
3. Stain overnight in GIEMSA. Krobock et al. speeded this up by
staining 1 hour in 1% Giemsa solution at 60° C.
4. Wipe slides. Remove excess blue by rinsing in 95% alcohol to which
a small amount of eosin has been added if necessary. Continue until
the collagen of the skin looks pink. Then dehydrate.

ORCEIN: dissolve 200 mg of Harleco’s synthetic Orcein in 100 mL of 70%
alcohol. Add 0.6 mL of concentrated HCl. The solution improves on
standing and has a long shelf life.
GIEMSA: one drop of stock in 20 mL of distilled water or pH 7.0
phosphate buffer.

RESULTS: Collagen is rose-pink, while elastic is dark brown to black.
Melanin is dark green to black. Bacteria and fungi are dark blue.

Looking at the book, I would suppose that Mehregan had just about
abandoned the use of this stain.

More information about the Histonet mailing list