[Histonet] coverslipping question

b427297 <@t> aol.com b427297 <@t> aol.com
Thu Nov 14 09:07:26 CST 2013

Are you using tape or glass coverslips?  We have found that when using tape coverslips, at least two very clean 100% alcohol steps must be added before slides go into the clearing reagent, be it xylene or Histoclear.  If not, brown spots appear on some tissues, usually liver and bone.  If there is any TRACE of water retained in the tissue, it will show up as a brown artifact on the tissue.  This doesn't happen with conventional glass coverslipping and mounting medium - I think the mounting medium compensates for the retained moisture in the tissues.

-----Original Message-----
From: Kim Donadio <one_angel_secret <@t> yahoo.com>
To: Gautier, Nicole M. <ngauti <@t> lsuhsc.edu>
Cc: histonet <histonet <@t> lists.utsouthwestern.edu>
Sent: Wed, Nov 13, 2013 1:37 pm
Subject: Re: [Histonet] coverslipping question

I would try adding 1 or 2 more 100% alcohols before the histoclear. 

Sent from my iPhone

On Nov 12, 2013, at 9:32 AM, "Gautier, Nicole M." <ngauti <@t> lsuhsc.edu> wrote:

> My lab has been having a problem with "specks" appearing on our slides after 
they come out of Histoclear.  Our protocol is to dehydrate in 2 minutes each of 
70%, 80%, 95%, and 100% ethanol before 2 minutes 2 times in Histoclear.  The 
slides are perfectly clear when they come out of the ethanols, but not when they 
come out of the Histoclear.
> Since it's not a problem we had when we first started, I tried changing out 
the Histoclear, but the problem remained.  At the end of last week, I changed 
out everything - all glassware, all ethanols, all Histoclear.  The problem went 
away and the 24 slides looked fine.  Today, the slides are looking speckled 
> So I guess my questions are: What could be causing the speckled appearance of 
the slides? and  How often should I have to change the ethanols and Histoclear?  
I seem to remember years ago when I did this that we only changed the solutions 
> Nicky Gautier
> Research Associate
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