Clontech and Molecular Probes recommendations for RE: [Histonet] sealing slides with diluted permount

[Gayle Callis]

Malcolm,

We found agarose and rubber cement to be terribly messy, and  since had 
conversations with Clontechs tech services on using diluted mounting 
media.  The same for Molecular Probes as their technical services one on 
one did recommend sealing the coverslip with Prolong Gold anitfade hard set 
- it does retract when dry, as we learned the hard way. They were also 
recommending the messy way.   Another reason for sealing is to keep the 
coverslip from sliding all over the place with potential for ruining a 
stained section (had it happen).

The only reason you can't use nail polish (per a Science publication on 
GFP) is that is contains isopropyl alcohol, and that is what ruins the GFP 
since the alcohol leaches into the aqueous mounting medias.  That is why we 
now use toluene or xylene based mounting medias since these two solvents 
are not miscible with water.

At 12:04 PM 12/19/2005, you wrote:
>I found this.
>http://www.clontech.com/clontech/techinfo/faqs/LivingColors/fixation.shtml
>
>Q What is the best method of sealing coverslips?
>
>A We recommend sealing coverslips with molten agarose or rubber cement. We 
>do not recommend using nail polish, because GFP is very sensitive to some 
>nail polishes. However, when using ProLong Antifade from Molecular Probes 
>(www.probes.com <http://www.probes.com/media/pis/mp07481.pdf> ) no further 
>sealing is necessary.
>
>
>Malcolm L. McCallum
>Assistant Professor
>Department of Biological Sciences
>Texas A&M University Texarkana
>2600 Robison Rd.
>Texarkana, TX 75501
>O: 1-903-233-3134
>H: 1-903-791-3843
>Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html
>
>
>________________________________
>
>From: histonet-bounces <@t> lists.utsouthwestern.edu on behalf of Caroline Bass
>Sent: Mon 12/19/2005 12:51 PM
>To: Histonet (E-mail)
>Subject: [Histonet] sealing slides with diluted permount
>
>
>
>Hello,
>
>I have some slides of GFP expressing tissue.  I am using the prolong
>gold mounting media from Molecular Probes to mount the coverslips but
>now I have to seal them.  I have read that the best way to do this is
>with diluted permount.  Unfortunately I have never sealed slides
>before and am having a hard time visualizing the process.
>
>What is a good starting dilution for the permount?  I was thinking
>1:10 with xylenes to start, but this is just a guess.  How exactly do
>you apply the permount?  Can you just pipet it around the edges or do
>you need to brush it on in some manner?  Does it have to be flush
>with the coverslip or can you have some buildup?
>
>Any advice would be appreciated.
>
>Thanks,
>
>Caroline Bass
>
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Gayle Callis
Research Histopathology Supervisor
Veterinary Molecular Biology
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610
406 994-6367
406 994-4303 (FAX)