I always used a charge or aplus slides on all of my paraffin-embedded
section. It's a 5uM thick. I did check the temperature after I boil the DAKO
TRS solution and it's 80-85C. Today I did the CD31, again and I had the TRS
at 75C and some edges around the tissue starting to lift off.
>From: "Amos Brooks"
>To: "Thu-Thao Pham" ,
>Subject: Re: Pretreatment.
>Date: Mon, 10 Dec 2001 19:57:41 -0500
> I'll bet it is your slides. Charged slides are a must. Also it would
>a good idea to keep a close eye on the temperature of the retrieval
>solution. If the slides are actually boiling in the solution, it is too hot
>and is unnecessarily agitating the slides. I would also caution against
>using float bath adhesives being used in conjunction with charged slides.
>The archives have countless explanations of why this goes wrong. Beyond
>that I'd look into the processing of the specimen.
>----- Original Message -----
>From: "Thu-Thao Pham"
>Sent: Monday, December 10, 2001 1:10 PM
> > Hi everyone,
> > As some of you may know that I post a questions regarding CD31 and
> > pretreated the slides with DAKO Target Retrieval Solution. However, my
> > tissue got eaten up and the mophology look back. Some tissues even about
> > come off the slides. I try to be very careful when I take the slides out
> > washing the slides.
> > I did as followed:
> > 1) Deparaffinized (Histoclear to water)
> > 2) Washed slides in TBST 3x for 5 min.
> > 3) Pretreatment with TRS by steam for 20 min.)
> > - first heat the TRS to boil and let the TRS
> > solution become clear before place slides into
> > the TRS heated solution (95C).
> > 4) Remove the jar from steamer and let it cool to room temperature for
> > 30min.
> > 5) Gently remove the slides and washed in ddH2O 3x for 5 min.
> > 6) Washed slides in TBST 3x for 5 min.
> > 7) Proceed with 3% H2O2 for 5min.
> > 8) Washed TBST
> > 9) CD31 diluted in 1% BSA (in TBST).
> > 10) Followed by DAKO kit
> > After preteated my slides can I leave at room temperature for at least
> > hour to cool down?
> > Many thanks if anyone can help.
> > Thao
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