Re: Thionin stain (Nissl, myelin, glia)

From:"J. A. Kiernan"

> Histo-Scientific Research Laboratories wrote:
> Does anyone recommend a thionin stain for NISSL bodies and
> glial cells?  We normally perform a LFB or Cresyl Violet for
> NISSL, but the researcher would like thionin.

Yes. Thionine is good for this purpose, preferable in my
opinion to cresyl violet. Cresyl violet, like thionine 
stains Nissl substance (cytoplasmic RNA of neurons) and 
also the DNA in all nuclei, including those of glial 
cells. LFB (luxol fast blue - it may be one of 3 or 4 
different dyes)stains myelin. 

LFB dyes are not always the ones of choice for myelin. 
There are easier methods using eriochrome cyanine R 
(= solochrome cyanine R = chromoxane cyanine R = CI 43820, 
Mordant blue 2), and there are more contrasty methods 
for myelin (black & white) using hematoxylin.

Myelin + Nissl on one slide looks pretty when done well
(needs microscope controlling, especially when LFB is
used). For many purposes, especially quantitative studies, 
it is easier to evaluate sections stained with either a 
Nissl or a myelin method than to work with a combination.
The combinations work best with thin (10-20 um or less)
sections. The thicker (50-100 um) sections commonly used
in neuro research work are not suitable for simultaneous 
staining of more than one abundant component of the 

In combined methods applied to paraffin or thinnish frozen
sections, a red Nissl stain gives better contrast to blue 
myelin than either thionine or cresyl violet. Neutral 
red serves well, after either a "luxol" blue dye or 
eriochrome cyanine R (CI 43820). It is usual not to
counterstain after one of the hematoxylin-based myelin
stains (black), because the gray matter and nuclei are
superbly transparent, except for fibers of passage that
can be seen and followed at higher magnification.

At Histo-Scientific Research Laboratories you surely
have access to published literature in this field,
so you'll know that there's a lot of it, and that it
isn't possible to prescribe a sure-fire recipe that
will stain myelin and/or Nissl ideally for every

This email repy to you and to Histonet is already too 
long, so I won't add a bibliography or my own anecdotal 
burblings (which would be 3 or 4 more screenfuls, in 
this matter of Nissl/myelin etc). If you need some
references I'll be happy to provide lots. Just ask,
but try to focus the question. 

Finally, it's thionine NOT thionin, despite wrong
spelling in catalogs, books etc. Ask for refs if you
don't believe me, and decide for yourself.
John A. Kiernan
Department of Anatomy and Cell Biology
The University of Western Ontario
London,   Canada   N6A 5C1

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