We don't have problems with our eosin after Xylene-free deparaffining. I use a 95% alcohol step just before the Eosin. Perhaps that might help. We don't air dry before coverslipping as my prof finds the quality not as good, so we use 100%IsOH before coverslipping; the mounting media you use will determine if it is ok to cover slip from IsOH. We just had a grad student compare Hot IsOH vs mineral oil vs Hot detergent as xylene free alternatives. I'll send you our final protocol if you like. Margaret Horne AVC, UPEI, "Gerard Spoelstra" <g.spoels...@murdoch.edu.au> 27/03/2013 11:06 AM >>> 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 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
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