AW: [Histonet] In Situ Hybridization Blue Background problem

Gudrun Lang gu.lang <@t>
Thu Jan 31 03:01:09 CST 2008

Just a thought. Do you dry your slides in an oven before staining?
We have noticed some strange effect with charged slides, when we did FISH on
cytopreps using charged slides. There was an enormous flourescent
background, when we let them just airdry. This background was diminished,
when we put them into the 60°C oven for half an hour. 
So I think, that the adhesive can be "destroyed" with heat, and therefore
decreases the chance for catching detection-reagens. 

It can also be a matter of insufficient washing with the vortex-stations.
When we had problems with patchy staining, the solutions was finally the
exchange of the complete slide-tableau to one with more vortex-stations.

Gudrun Lang
Biomed. Analytikerin
Akh Linz
Krankenhausstr. 9
4020 Linz

-----Ursprüngliche Nachricht-----
Von: histonet-bounces <@t>
[mailto:histonet-bounces <@t>] Im Auftrag von Brandon
Gesendet: Mittwoch, 30. Jänner 2008 17:13
An: histonet <@t>
Betreff: [Histonet] In Situ Hybridization Blue Background problem

I have an In Situ Hybridization question for you guys and gals. I'm
currently performing In Situ HPV and EBV tests using an automated machine
called the XT by Ventana. The tissue is parrifin embedded, cut at 4 microns
and place on charged slides. NBT/ BCIP is used as our chromogen. I am
beginning to notice some background staining that varies in intensity. When
it occurs on our QC slide, it can be strong enough to fail a run. The
background resembles a "blue haze" that can cover the tissue making
interpretation difficult. At other times the blue haze surrounds the tissue
rather than covering it which makes me believe that there could be some sort
of hydrophobic or other chemical rxn taking place to shield the tissue. I'm
not certain if these two problems are related, but both occur completely
random and always vary in intensity. If I can find the culprit of this
problem it would lead to cleaner slides and more conclusive resulting. If
anybody can help it would be much appreciated. Thanks   P.S.  Ventana has
been notified of the problem and engineers have been sent multiple times to
try to figure it out.
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