Re: 10% NBF for TEM fixation?

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From:"R.Wadley" <> (by way of Marvin Hanna)
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	Dear Karen,

	Bouin's would work for EM in the context of this discussion, ie,

	Picric Acid in electron microscopy is a useful fat fixative instead of
OsO4, particularly if you wnat to do immuno.  OsO4 is no good for immuno
because it blocks/alters antigen binding sites, but picric acid does not
but still gives reasonable lipid fixation.

	So, Bouin's fixed Zebra fish, processed (via your normal protocol) may
give that ideal compromise between rapid fixation and good fixation that
gives the ideal EM images.

	As to adipose tissue effecting penetration, Yes.  formaldehyde/glut
fixatives are generally water based & therefore will have a tougher time
penetrating a lipid barrier.  Ethanol based fixatives will get into the
tissue at a better rate.


	Rob W.

At 10:31 01/20/2000 +0000, you wrote:
>Carla & John,
>Here's some zebrafish-specific information that may affect your decisions
on what to
>do next:
>The zebrafish very often has a thick layer of adipose tissue right under
it's skin.
>My presumption is that this may affect the penetration of your postfix.
When I do
>paraffin sections of the fish, I use Bouin's or an alcohol-based formalin
>(Davidson's or Dietrich's), presumably to increase penetration.  Of course
>fixes are not useful for EM.  If John or the other experts think the
penetration will
>be affected by this layer of adipose tissue, you may want to skin the fish
before you
>post fix.  It's relatively easy to skin a fixed zebrafish.  I just make a
few cuts in
>the skin at the back of the head, and use forceps to peel off the skin and
>layer of adipose tissue.
>Will this adipose tissue affect the penetration?
>Karen in Oregon

R. Wadley, B.App.Sc. M.L.S, Grad.Dip.Sc.MM
Laboratory Manager
Cellular Analysis Facility
School of Microbiology & Immunology
UNSW, New South Wales, Australia, 2052
Ph (BH) 	+61 (2) 9385 3517
Ph (AH)	+61 (2) 9555 1239
Fax 	+61 (2) 9385 1591

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