Re: How long are subbed slides good?

From:Lorraine Gibbs

I can only chirp in with more of the same. For "trouble-prone sections" such
as thick brain sections (40um-100um) we mount floating sections from Tris
buffer, dry tilted not flat, following air-dry dry overnight at 37C.
We use gelatin subbed slides (gelatin=1.0% chrome alum=.07%).
For Cresyl, start the slides in 100% EtOH x 2 for 30-40 minutes for thick
sections. We then defat in Xylene for the same amount of time, finally
sections are hydrated and stained in the usual way.
Good luck!

Lorraine Gibbs
Physiology & Biophysics
University of Washington
----- Original Message -----
From: "Frouwke Kuijpers" 
To: "Melissa Jensen" ; ;
"Cathy Gorrie" 
Sent: Friday, April 12, 2002 6:55 AM
Subject: Re: How long are subbed slides good?


I had the same trouble with 40 um ratbrainsections and a cresylviolet
staining.
What I did succesfully : after mounting the slices on the glass (coated with
0,5% gelatin and 0,05% chroomaluin) : overcoat with a 0,5 % gelatin
solution, by soaking the slides 10 sec, than airdry. Impregnation time
during the staining is a little bit longer.
I don't know what influence it has on other stainings.

Frouwke

F.J.Kuijpers-Kwant
Dept. Cellular Animal Physiology
University of Nijmegen
Toernooiveld 1
6525 ED Nijmegen
frouwke@sci.kun.nl

----- Original Message -----
From: "Melissa Jensen" 
To: ; "Cathy Gorrie" 
Sent: Friday, April 12, 2002 3:06 AM
Subject: Re: How long are subbed slides good?


Maybe using gelatin in your bath water is the problem.I worked in a histo
lab prior to the one I am at.We only used gelatin there..Had trouble keeping
tissue on slides during gms procedure etc.I was introduced to Stay on at my
current facility.It is great! Nothing falls off..no curling,You don't even
have to let slides cool before putting them in the first xylene....There is
another product called Halt.We tried it..but it reduced the surface tension
of the water so much..All tissue no matter what curled...didn't adhere to
the slide......We use subbed slides..they all stay in tact..
Hope this helps!
----- Original Message -----
From: "Cathy Gorrie" 
To: 
Sent: Thursday, April 11, 2002 7:43 PM
Subject: Re: How long are subbed slides good?


> We regularly cut 30-40 Ám rat brain sections, place on subbed slides
> (0.8% gelatin with 0.05% chromium potassium sulphate) and air dry
> overnight before staining. Normally this is absolutely fine,and the
> age of subbed slides has varied from a few weeks to very old.
>
> Recently however, we have been having trouble with the same problems
> you are experiencing, BUT only on one series and one type of
> staining, rather than just any liquid. ie sections have been cut and
> placed on slides in series of two, one stays on through the stain
> over 4 days but the Cresyl Violet (10 mins) stained sections are
> lifting. I don't think it is the age of the subbing therefore, but
> possibly is due to the quality of subbing or staining solutions.
> There was some discussion on this list a while ago about alkaline
> solutions causing gelatin to become more soluble in water, hence the
> addition of chrome alum etc etc, and I suspect this is the root of
> the problem. Maybe one batch of subbing solution was not made
> correctly, or the a batch of staining solution is slightly more
> alkaline than usual.
>
> Hope this gives you a starting point.
>
> Cathy
>
>
> At 2:37 PM -0400 10/4/02, Knight, Brandon wrote:
> >I've been having a problem with 40um mouse brain sections coming off of
> >subbed slides when placed in any liquid (for instance, when going through
> >the staining process).  The edges may fold over, they may wrinkle, or
slide
> >off completely.  They were sufficiently dried (some for a couple of
weeks).
> >The slides that I'm using were subbed in August of 2000.  Is it possible
> >that the gelatin has degraded and that these slides are no longer subbed,
or
> >could there be another explanation?
> >
> >Brandon Knight
> >Children's Research Institute
> >Center for Genetic Medicine
> >Children's National Medical Center
> >Washington, D.C.
>
> --
> ---------------------------------------------------
> Cathy Gorrie
> Scientific Officer
> Neural Injury Research Unit,
> School of Medical Sciences,
> University of New South Wales
> Sydney, N.S.W. 2052
>
> Phone: 61-2-9385 2462
> Fax   : 61-2-9313 6252
> e-mail: c.gorrie@unsw.edu.au
>
>












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