Re: Washed Out Nuclei
| From: | RichardWHorobin@aol.com |
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Good morning folks - Well here in Scotland its 7.45am, blue skies, snow on
the neighbouring mountain, butnone down here - - Apologies to the
geographically less fortunate - -
Ann asked:
> Can anyone tell me why, intermittently, we are having slides that appear to
> have washed out nuclear staining. Mostly on endometrium, sometimes on
> reprocessed skin. Everything is processed/stained/etc. together. Does
> anyone have a good reprocessing schedule?
And Dana suggested she thought this sort of problem was linked to 'old'
xylene.
Well, it could be ineffective dewaxing. Secion dewaxing is certainly not
uniform. For instance the wax often comes out of nculei more slowly than form
surrounding cytoplasms. And not all nuclei dewax at the same rate. Obviously
if dewaxing is 'overall effective' then such differential effects are
irrelevant. But if the dewaxing is loncomplete, then some regions may be
more complete than others - and waxy nuclei do not stain strongly, the wax
excluded the dye.
And yes, damp xylene (eg end-of-day xylene) might well reduce the efficacy of
the dewaxing process.
I came across this effect when the factor limiting the efficacy of dewaxing
was low temperature. In a poorly heated lab, in the winter, putting cold
paraffin sections into cold xyelene resulted in slow dewaxing. I do remember
we solved this by warming up the xylene - we were manually processing, so
maybe put the xylene dish on the warmed slide-mounting tray?
Aside 1 - If you did this, dont tell the safety officer (I WAS the SO - - ).
Aside 2 - It was Dr Dan Goldstein - who died last year - who did the
trouble-shooting, if I remember correctly.
Well, try fresh / not-cool xylene and see! Good luck, Richard Horobin
Institute of Biomedical & Life Sciences, University of Glasgow
T direct 01796-474 480 --- E RichardWHorobin@aol.com
"What should we expect? Everything."
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<HTML><FONT FACE=arial,helvetica><FONT SIZE=2>Good morning folks - Well here in Scotland its 7.45am, blue skies, snow on
<BR>the neighbouring mountain, butnone down here - - Apologies to the
<BR>geographically less fortunate - -
<BR>
<BR>Ann asked:
<BR>
<BR><BLOCKQUOTE TYPE=CITE style="BORDER-LEFT: #0000ff 2px solid; MARGIN-LEFT: 5px; MARGIN-RIGHT: 0px; PADDING-LEFT: 5px">Can anyone tell me why, intermittently, we are having slides that appear to
<BR>have washed out nuclear staining. Mostly on endometrium, sometimes on
<BR>reprocessed skin. Everything is processed/stained/etc. together. Does
<BR>anyone have a good reprocessing schedule? </BLOCKQUOTE>
<BR>
<BR>And Dana suggested she thought this sort of problem was linked to 'old'
<BR>xylene.
<BR>
<BR>Well, it could be ineffective dewaxing. Secion dewaxing is certainly not
<BR>uniform. For instance the wax often comes out of nculei more slowly than form
<BR>surrounding cytoplasms. And not all nuclei dewax at the same rate. Obviously
<BR>if dewaxing is 'overall effective' then such differential effects are
<BR>irrelevant. But if the dewaxing is loncomplete, then some regions may be
<BR>more complete than others - and waxy nuclei do not stain strongly, the wax
<BR>excluded the dye.
<BR>
<BR>And yes, damp xylene (eg end-of-day xylene) might well reduce the efficacy of
<BR>the dewaxing process.
<BR>
<BR>I came across this effect when the factor limiting the efficacy of dewaxing
<BR>was low temperature. In a poorly heated lab, in the winter, putting cold
<BR>paraffin sections into cold xyelene resulted in slow dewaxing. I do remember
<BR>we solved this by warming up the xylene - we were manually processing, so
<BR>maybe put the xylene dish on the warmed slide-mounting tray?
<BR>
<BR>Aside 1 - If you did this, dont tell the safety officer (I WAS the SO - - ).
<BR>Aside 2 - It was Dr Dan Goldstein - who died last year - who did the
<BR>trouble-shooting, if I remember correctly.
<BR>
<BR>Well, try fresh / not-cool xylene and see! Good luck, Richard Horobin
<BR>
<BR>
<BR>Institute of Biomedical & Life Sciences, University of Glasgow
<BR><B>T direct 01796-474 480 --- E RichardWHorobin@aol.com</B>
<BR><I>"What should we expect? Everything."</I></FONT></HTML>
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