[Histonet] Solochrome Cyanine R

[John Kiernan]

First, the name! In most catalogs this dye is
called eriochrome cyanine R, so that's the name
used in the current (10th) edition of Conn's
Biological Stains. Two synonyms, much used in
the literature, are solochrome cyanine R (ICI's 
trade-name, I think), and chromoxane cyanine R
(which originated, I think, with some pre-war
German company and was used in the 9th edn of Conn's). 

The Colour Index name is CI Mordant blue 3; CI 43820.

The dye has several uses in histology and there
are several published procedures for staining
myelin. My favourite is as follows:

__________________________________________________________________

Iron-eriochrome cyanine R for myelin
(after Kiernan, 1984; original was Page, 1965).

Staining solution.

0.21 M aqueous ferric chloride (5.6% w/v FeCl3.6H2O):  20.0
ml
Eriochrome cyanine R (C.I. 43820):                      1.0
g
Concentrated (95-98% w/w) sulphuric acid:               2.5
ml
Water:                                         to make  500
ml

(The ingredients take 2-5 minutes to dissolve, with
stirring. Filtration should not be necessary. The solution
can be kept and used repeatedly for at least 8 years.) 

Differentiating solution for myelin.

Any one of the following aqueous solutions may be used. See
also Note below.

Iron alum (NH4Fe(SO4)2.12H2O):     10% w/v
Ferric chloride (FeCl3.6H2O):     5.6% w/v
Ferric nitrate (Fe(N03)3.6H2O):   7.3% w/v

(The differentiating solution keeps for a few years, but may
be used only once. Discard if it is cloudy or if there is a
thick layer of pale insoluble material in the bottom of the
bottle.)

Counterstain.

After a myelin stain, use a red basic dye (0.5% aqueous
neutral red or safranine is suitable) to stain nuclei and
Nissl substance.

Method (paraffin sections of formaldehyde-fixed animal
tissues, especially CNS).

1. Stain hydrated sections for 15-20 minutes are needed.
(The slides may remain in the dye solution for 30 minutes
without harm.) 
2. Wash in running tap water, 30 seconds, or in 3 changes of
distilled water. This is to remove unbound dye.
3. Immerse in the differentiating solution until only the
myelin (white matter of CNS) retains the stain. This usually
takes 5-10 minutes. It is sometimes impossible to decolorize
the nuclei completely without losing some intensity in
myelin.
4. Wash in tap water (running, or three or four changes) for
about 5 minutes.
5. Apply a counterstain, as desired.
6. Dehydrate, clear, and cover using a resinous mounting
medium.

Result.

Myelin and red blood cells dark blue.
________

Note on differentiation. Clark (1979) recommended an
alkaline differentiating solution for myelin staining: a
freshly prepared 1% (v/v) dilution of ammonium hydroxide.
This acts in a few seconds, and it is easy to remove too
much of the blue dye-metal complex. 
________

References.
Clark, G. (1979). Staining with chromoxane cyanine R. Stain
Technology 54: 337-344.
Kiernan, J.A. (1984). Chromoxane cyanine R. II. Staining of
animal tissues by the dye and its iron complexes. Journal of
Microscopy 134: 25-39.
Page, K.M. (1965). A stain for myelin using solochrome
cyanin. Journal of Medical Laboratory Technology 22:
224-225.
_________

The colour is darker than myelin that has been stained 
with a luxol fast blue. If the differentiation (Step 3)
is done instead with acid-alcohol, the result is a clean
blue nuclear stain similar to that seen with a haemalum
but more resistant to extraction by subsequently applied 
acidic counterstains. 
____________________________________________________________

--
-------------------------------
John A. Kiernan
Department of Anatomy and Cell Biology
The University of Western Ontario
London,   Canada   N6A 5C1
   kiernan[AT]uwo.ca
   http://publish.uwo.ca/~jkiernan/
   http://instruct.uwo.ca/anatomy/530/index.htm
_______________________________
"Ant S." wrote:
> 
>    Does anyone out there know about Solochrome Cyanide R stain for myelin
>    staining?  Please tell me where we could find out about the stain, and
>    if  you  have  any  tips  or recipes  you  could share,  that would be
>    super nifty.
> 
>    Your help is appreciated, thanks.
>    Antoinette Swensson
>    Univeristy of Washington/Harborview Medical Center
>    Neuropathology
>      _________________________________________________________________
>