[Histonet] IF-fading retardants

Kim Merriam kmerriam2003 <@t> yahoo.com
Thu Aug 30 08:08:25 CDT 2007


Are you saying that IF stained slides can go through the normal dehydration/clearing process and be mounted in a xylene-based mounting medium?

Kim Merriam, MA, HT(ASCP)
Cambridge, MA

----- Original Message ----
From: John Kiernan <jkiernan <@t> uwo.ca>
To: AGrobe2555 <@t> aol.com
Cc: histonet <@t> lists.utsouthwestern.edu
Sent: Wednesday, August 29, 2007 1:31:52 AM
Subject: Re: Re:[Histonet] IF-fading retardants

Why do you "suspect you would kill your signal" by dehydrating, clearing and  mounting a slide carrying a fluorescently labelled antibody? 

Alcohol coagulates proteins on-the-spot, complete with any covalently bound fluorescent tags. This was established for lectin histochemistry 30 years ago, and has been well documented for fluorescently labelled antibodies in more recent years. The intensity of  fluorescence emission may be less in DPX than in buffered glycerol, but who has done comparisons? 

A fairly recent study strongly favours alcohol dehydration, clearing and mounting in a non-flourescent  recinous medium. for permanent immunofluorescence slides.

John Kiernan
Anatomy, UWO
London, Canada
----- Original Message -----
From: AGrobe2555 <@t> aol.com
Date: Monday, August 27, 2007 14:31
Subject: Re:[Histonet] IF-fading retardants
To: histonet <@t> lists.utsouthwestern.edu

> Carl,
> I used to use "FluorSave" mounting medium from Calbiochem (Cat 
> #  345789).  
> This mounting medium "hardens" so the coverslip doesn't 
> move  and you don't 
> have to seal around the edges with nail polish.  It also 
> kept  the signal from 
> fading.  I still stored the slides horizontally at 4C 
> or  -20C.  I wouldn't 
> suggest try in to dehydrate through xylene and  
> coverslipping, as I suspect you 
> would kill your signal.  
> Albert
> Albert C.  Grobe, PhD
> International Heart Institute of Montana Foundation
> Tissue  Engineering Lab, Saint Patrick  Hospital
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