[Histonet] Eosin not working for xylene free H&E

[Rene J Buesa]

Totally xylene free staining is a consistent sequence described as:
1- dewax the sections with 2 washes of liquid dish washer soap (any brand will do) in a 2% aq. sol. at 95ºC of 1 minute each.
2- 2 consecutive 1 min. each wash with water at 90ºC and to water at RT
3- stain as usual, any routine or special stains → to pure ethanol
4- the stained sections are placed in an oven at 65ºC for 5 minutes
5- cover as usual.
No problems with the staining
For more details please go to http://www.histosearch.com/rene.html and read the "Xylene free" article.
René J.
 

From: Gerard Spoelstra <G.Spoelstra <@t> murdoch.edu.au>
To: histonet <@t> lists.utsouthwestern.edu 
Sent: Wednesday, March 27, 2013 10:06 AM
Subject: [Histonet] Eosin not working for xylene free H&E

Hi everyone,

I have not had any success optimising the eosin counterstain in the absence of xylene. We routinely use 0.5% eosin Y in 95% ethanol. I've tried taking sections through to this to absolute ethanol but then air drying , but it leaves blotches of eosin.  Using eosin 0.5% aqueous with a few drops of acetic acid doesn't appear to work, the keratin stains heavily but not elastin. After washing in water I leave for a short time in the oven then at room temperature for 5 minutes then coverslip with the dako atomatic cover slipper.  For all the laboratories that have gone xylene free, are you just putting up with less than optimal eosin staining?

Gerard Spoelstra  
Murdoch University
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