[Histonet] RE: brain sections falling off slides during IHC

[Linke, Noelle]

Hi Christina,

In my experience, there are a few things to try.  First, don't soak your blocks too long before cutting, and try not to let too much water get underneath the sections as you pick them up from the waterbath.  I wouldn't put them in an oven at all, just let them air dry as long as you can (overnight, a day if you can spare the time) before staining.

If you're having troubles, I wouldn't use the autostainer at all (it rinses too aggressively for some tissues), I would do them manually and never let the stream from your wash bottle touch the tissue, gently rinse after each step.

I hope this helps.

Noelle

Noёlle Linke M.S., HTL(ASCP)QIHC
Manager, Histology Services
Department of Pathology & Laboratory Medicine
David Geffen School of Medicine at UCLA
Phone: 310-825-7397
Pager: 97471
nlinke <@t> mednet.ucla.edu

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Thurby, Christina
Sent: Wednesday, February 11, 2009 7:38 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] brain sections falling off slides during IHC

Hello all,
Can any of you all with experience immunostaining brain sections please help.  My sections are almost completely washing off the slides.  I am using 10% NBF fixed canine brain sections.  Embedded in paraffin and sectioned at 4 microns on plus slides with distilled water in the water bath.  The slides are dried at 65 C for 15 minutes and allowed to stand overnight.  I am using a Dako Autostainer for staining.  There is no antigen retrieval step.  My staining procedure is approximately 3.5 hours long.  At the completion of the procedure, the tissue on my slides is 50-80% washed off.

Any thoughts??
Christina Thurby
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