[Histonet] H&E question

Bryan Llewellyn llewllew <@t> shaw.ca
Wed Jan 9 15:47:35 CST 2008

Chances are the biopsies are too dark and mucin is staining because you are 
using Gill III.  This is a very strong hemalum with 6 grams hematoxylin per 
litre.  If all is oxidised I would be surprised if mucin did not stain. 
Strong hemalums which are fully oxidised nearly always stain acid mucins, 
especially if they are becoming acid deficient.

Try switching to Gill II or another hemalum with less hematoxylin, or dilute 
your Gill III by half or by a third with a solution made up the same as the 
Gill III, but without the hematoxylin and oxidiser.  Try Cole's progressive 
hemalum, or some other type.  Cut a lot of sections and carry out some 
experiments on typical types of tissues you get until the staining is 

Go to http://stainsfile.info and read the articles there about formulating 
hemalum and the other articles about hematoxylin staining.

Rather than use deionised water, store tap water over some marble chips in a 
20 gallon container and use that for the final rinses after blueing the 
hemalum and before the eosin.  Calcium ions are reported to improve eosin 
staining.  They also would make the water alkaline and ensure the blueing is 

Bryan Llewellyn

----- Original Message ----- 
From: "Sharon.Davis-Devine" <Sharon.Davis-Devine <@t> carle.com>
To: <histonet <@t> lists.utsouthwestern.edu>
Sent: Wednesday, January 09, 2008 11:08 AM
Subject: [Histonet] H & E stain question

We are continually having a problem with our H & E stain.  We currently
use Gill III Hematoxlyin on a staining machine with a running water
bath.  Our pathologists do not like to have the pH of the Heme higher
than 2.5 because it makes the small biopsy sections too dark and the
mucin appears blue.  My question to all of you is, has any of you
experienced this issue and if so, how did you go about getting it fixed?
I had suggested switching to a different heme such as Harris and maybe
switching our running water bath to DI water.  We have significant
flooding in our area and there will be more bleach in the city water
supply.  Does any of this sound feasible?  Help please.  Thanks.

Sharon Davis-Devine, CT (ASCP)

Cytology Supervisor

Carle Clinic

602 West University

Urbana, Illinois 61801

Phone:  217-383-3572

Email:  sharon.davis-devine <@t> carle.com

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