Re: Need help with GMA sectioning

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From:Karen S Pawlowski <>
To:"Jacqueline F. Webb" <>
Date:Thu, 5 Aug 1999 09:03:07 -0500 (CDT)
Content-Type:TEXT/PLAIN; charset=US-ASCII


I do alot of section of large (2 - 4 cm X 2 cm) blocks embedded in JB-4
and this has always been a problem that had to be concidered.  I am
usually embedding inner ear tissue of rodents and the wrinkling in my
case occurs between the inner-ear fluid space and the surrounding
decalcified bone.  What is happening is that when the GMA is exposed to
water in the water bath, it rapidly adsobes water, which causes it to
distend.  Since it is the GMA itself that is picking up water, uneven
adsorption and distention occurs between the dense tissue and the fluid
spaces.  A softer block usually reduces this effect, as there is already
some water in the softer blocks.  There are some tricks you can try to
improve your sections right now:

I use amonia in the water bath (10 ml strong amonia in 3 L distilled
water) and I heat the water bath on the slide warmer that I use to dry the
slides.  I also "fog" the sections before floating them on the water- in
order to partially rehydrate them. Again, this reduces the amount of water
that is rapidly adsorbed when the section hits the water. You could also
try rehydrating the blocks slightly by placing them in a humidity chamber
over night.  Good luck.

Karen Pawlowski, MS
Sr. Res. Assoc./UT Southwestern Med. Ctr.
PhD Candidate/UT Dallas
Dallas, TX  

On Wed, 4 Aug 1999, Jacqueline F. Webb wrote:

> Hope someone out there can help.
> We have been sectioning fish heads (portions of large ones or whole small
> ones) for a study of sensory systems and swim bladder specializations in
> butterflyfish.  These fish have a very thick stratum compactum in their
> dermis, which is composed of fibrous collagen.  Infiltration has been good
> (over 2 days), translucent tissue is the result. We also use a bit of
> vacuum to get rid of bubbles. Polymerization is carried out in the frig for
> larger blocks because of rising temperatures.  Sections at 5 um are fine,
> but when we float them out on water (room temp or very warm) the plastic
> stretches, the internal soft tissues stretch, but the collagen in the
> dermis doesnt stretch. The result is ruffles radiating out from the tissue
> and folds in the internal tissues. Staining and subsequent drying does not
> change this situation.  This prevents any decent publishable photos from
> being taken at lower magnifications (which is essential).  The tissue (a
> marine fish) was fixed in 10% formalin in Seawater or in phosphate buffer
> and decalcified (with radiographic confirmation) by Cal-Ex (Fisher)
> overnight with 3-4 hours running tap water rinse.
> HELP??
> Jackie Webb
> Asst. Prof. Biology
> Villanova University
> Jacqueline F. Webb
> Department of Biology
> Villanova University
> Villanova, PA  19085
> 610-519-7086 (office/lab)
> 610-519-7863 (FAX)
> If you do not receive a reply to a message, or
> if your message is bounced, please send the message
> to me at:

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