Kari,

As Aidan says, if the mounting media is too thick, you could have
trouble with focus planes. It can also be caused by sections which are
too thick or wrinkled sections, or water bubbles trapped under the
section. When drying the slides, be sure that not only is the paraffin
melted on the slide, but that all the water is gone too.

The cloudiness on your slides could be caused by water on the slide.
Check to make sure your 100 percent alcohol is really 100 percent and
not contaminated with water. Again, make sure the water is gone before
starting staining.

I haven't heard of any mounting media that is incompatible with xylene,
but it is a possibility and could cause cloudiness (there are some
mounting media's which are incompatible with various xylene
substitutes).

The holes in you blocks are almost certainly caused by rushing your
embedding. They are caused by air bubbles trapped in the mold. Allowing
the paraffin to cool too much before adding extra to top-up the mold
will cause a weak join-line around the tissue in the block and that will
cause cracking when sectioning. You need to be sure the paraffin is
still warm before adding the extra to fill the mold. In a well-embedded
block you should not see any line between the inital paraffin put in the
mold and the extra used to top it up.

Tim Morken, B.A., EMT(MSA), HTL(ASCP)
Infectious Disease Pathology
Centers for Disease Control
MS-G32
1600 Clifton Rd.
Atlanta, GA 30333
USA

email: tim9@cdc.gov
       timcdc@hotmail.com

FAX:  (404)639-3043




----Original Message Follows----
Date: Fri, 26 Feb 1999 15:01:31 +1300
From: "Aidan C. Schurr" <aidan.schurr@stonebow.otago.ac.nz>
Subject: Re: Slide "Plains"
To: ODDBALSTER@aol.com
Cc: HistoNet <HistoNet@Pathology.swmed.edu>

Kari,
Just a couple of suggestions - the different planes you describe are
quite
possibly due to different thicknesses of the mountant under the
coverslip.
Too much mountant leads to a different refractive index, which can give
the
problem you describe.  Not too sure with the haziness - could be the
same
problem if the mountant layer is **very** thick, but this seems
unlikely.
Another possibilty is that you have water in your zylene, or dehydration
has been incomplete.  You could perhaps try using two changes of xylene.

Hope this is of some help

Good luck
Aidan, New Zealand


>Hello Histonetters, I was hoping you could help me a couple problems.My
>pathologist was complaining that the stained H & E sections in our
daily work
>was somewhat on different "plains" and needed constant focusing while
under
>the scope. I sat down with her and went over some slides, these
sections were
>somewhat "fuzzy or hazy" in some places and crisp in other, which makes
me
>think possibly it's the mounting medium...we've been using this medium
for
>sometime (resinous) and unfortunately we're on contract so my options
are
>limited (for purchasing that is). Could it possibly be the
deparaffination
>process? We microwave our sections to melt the paraffin, and put them
in
>Xylene for 6 minutes to clear before sending them down the stainer. It
does
>not happen on all the tissue, and there does not seem to be any
preference as
>to type of specimen either (happens to bx's and surgicals). Any
suggestions?
>Also, we seem to be getting holes in our blocks after embedding, the
VERY
>irritating experience is becoming more and more frequent. We have fresh
>paraffin in our dispenser (and processor) and have a cold plate at
optimal
>temperature, yet it seems at least 4-5 blocks per day are "breaking"
when
>trimming because of these holes. Is it possible that we may be
embedding too
>fast? Not giving the warm paraffin time to combine with the cold
paraffin
>after inserting the tissue in the mold? Could it be that we aren't
cleaning
>the molds enough? Again, any suggestions? Thanks for your help in
advance, man
>I love this net......
>
>Kari Zajic HT,MLT
>Lead Histotech
>Florida


/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/
	Aidan C. Schurr  BMedLabSc
	   Histology Service Unit		++64 3 4797152
	      Pathology Department
	         Dunedin School of Medicine

	            acschurr@bigfoot.com
   	               aidan.schurr@stonebow.otago.ac.nz
\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\/\








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