[Histonet] Bone IHC

Adam . anonwums1 <@t> gmail.com
Thu Oct 7 15:08:29 CDT 2010

I've been experimenting with different ways to solve this problem myself. I
fix my tissues in 10% zinc buffered formalin and decalcify in formic acid
for 72 hours, followed by embedding and cutting 5 uM sections.

>From trial and error, I've determined that incubating the slides flat on a
slide warmer at 37C (or in my case, the bottom of an unused bacterial
incubator) can prevent nearly all the detachment you observe but it's time
dependent. I think the problem is that the sections get small amounts of
water underneath them when you scoop them up off the water surface during
sectioning and during HIER, that water boils and shears off the slide.

If you leave the slides overnight, the slides were get destroyed during
HIER. However, if you leave them for a week, they tend to be nearly
untouched even at 95C for 10 mins. I'm currently in the process of
determining if a few days is enough time.


On Thu, Oct 7, 2010 at 2:20 PM, Liz Chlipala <liz <@t> premierlab.com> wrote:

> We lower the temp of retrieval to 70C for 2 hours and have good success
> with that.
> Liz
> Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
> Manager
> Premier Laboratory, LLC
> PO Box 18592
> Boulder, Colorado 80308
> office (303) 682-3949
> fax (303) 682-9060
> www.premierlab.com
> Ship to Address:
> 1567 Skyway Drive, Unit E
> Longmont, Colorado 80504
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Vanessa
> J. Phelan
> Sent: Thursday, October 07, 2010 1:14 PM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] Bone IHC
> Hi everyone,
> I was wondering if anyone has any tricks on how to get bone sections to
> stop lifting off the slide through the IHC process? I leave them in the
> oven for quite a while to make sure they are baked on, however after
> antigen retrieval  (pressure cooker for 20mins) most of the boney part
> of the tissue comes off and the marrow and muscle stays put! The
> sections are cut onto superfrost plus slides.
> Any help would be much appreciated, thanks.
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