Re: [Histonet] staining mast cells and eosinophils

From:John Kiernan

Yes, it's easy to stain both cell-types. First stain with eosin at a high pH (such as 8) to obtain selective coloration of eosinophils. (Paneth cell granules will also stain if your tissue is intestine.) Wash in slightly acidified water, then stain with dilute toluidine blue at a low pH (less than 2 to be selective for mast cells, cartilage matrix and some types of mucus). The staining is metachromatic - purple to red, rather than blue, but not likely to be confused with the eosin colour. Rinse in slightly acidified water, dehydrate in 3 changes of 100% alcohol, clear in xylene and apply coverslip.
Slightly acidified water = approx 0.5% acetic acid. Check with a microscope at both the washing stages to make sure the eosinophils and mast cells are adequately stained. As described above, nuclei should be unstained. If you want blue nuclei, use the toluidine blue at pH 3 to 4, being careful to avoid over-staining.

John Kiernan
Anatomy, UWO
London, Canada.
----- Original Message -----
From: Moran Elishmereni 
Date: Tuesday, August 28, 2007 5:36
Subject: [Histonet] staining mast cells and eosinophils

> Hello,
> I am very new at histology and immunohistochemistry, so please 
> excuse the
> simplicity of my questions. I wish to stain parrafin-embedded slides
> (murine/human skin and lung) for mast cells and eosinophils. 
> That is- I want
> to be able to detect both cells on one slide. Is there anyway to 
> stain with
> toluidine blue and also counterstain with another dye for eosinophils?
> Alternatively- can i stain for toluidine blue to get mast cells, 
> and then do
> (on the same slide) immunohistochemistry using an anti-MBP 
> antibody to
> detect eosinophils? I guess its more of a general question- can 
> one use a
> simple dye AND immunohistochemistry (with antibodies) on the 
> same slide, or
> is there interference? I would be very glad to receive advice 
> and protocols.
> Many thanks,
> Moran Elishmereni
> Department of Pharmacology
> School of Pharmacy, Faculty of Medicine
> The Hebrew University of Jerusalem
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