I'm looking to contact Rebecca (Becky) Orr. Does anyone know her current email
address? The one I have bounced back.
Thanks, Andrea
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Hi All, Do you have a protocol for formalin fixation (and processing to FFPE)
of a previously snap frozen piece of tissue? Or advice as to the best method of
snap freezing for such a process (assume isopentane slurry)? We have done this
for several tissues with our standard fixation and
Congratulations Dr. Adam on your recent defense (yay). Thanks for the kind
words, I have enjoyed emailing with you and discussing bone IHC. We will miss
you on Histonet and I certainly hope you return to research once you have
completed your clinical years (if not sooner).
Best, Andrea
Hi All,
I am wondering if anyone knows of any particular publications or studies that
have examined manners in which one can predict whether an antibody in a large
panel may work in IHC. Let's say you are faced with 50-100 Abs and you need to
determine which works in IHC, but all you know is
Hi All,
I am doing a survey and will be happy to compile results and share if folks
will respond! What is your favorite antigen retrieval method and/or panel?
Buffer
Source/composition
Temperature
Device
Thanks, Andrea
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Agree, switching to Leica with disposable blade holder changed my life. Never
looked back. Love the CM3050S.
--- On Sun, 7/10/11, Rene J Buesa rjbu...@yahoo.com wrote:
From: Rene J Buesa rjbu...@yahoo.com
Subject: Re: [Histonet] In search of a cryostat...
To: histonet@lists.utsouthwestern.edu
Use the TUNEL assay, several very easy to use kits available. Roche's Cell
Death Detection kit with FITC might be the easiest.
--- On Thu, 4/1/10, Alexandra Meinl alexandra.me...@gmail.com wrote:
From: Alexandra Meinl alexandra.me...@gmail.com
Subject: [Histonet] Assay for cell death needed
Very interesting! Coming out the nose is definitely bad for any work I have
done in the past - lungs get blown out, liver doesn't perfuse well and bone
marrow looks horrific. However, if you are working with PFA and doing a
post-fix anyway, you will probably be fine. If you are using GA and
We used to do overnight in our DAKO Autostainer many years ago, and we kept
many containers of dH20 in the incubator to make a humid chamber. Seemed to
work well - although we realized overnight was overkill and 1-2 h was
sufficient. In fact, given this we started to always put humid chambers
else,
Andrea
--- On Wed, 3/3/10, Mauricio Avigdor bitesizell...@gmail.com wrote:
From: Mauricio Avigdor bitesizell...@gmail.com
Subject: Re: [Histonet] IF staining on peritoneal macrophages
To: Andrea T. Hooper andreahoo...@rocketmail.com,
histonet@lists.utsouthwestern.edu
Date: Wednesday
macrophages
To: Andrea T. Hooper andreahoo...@rocketmail.com,
histonet@lists.utsouthwestern.edu
Date: Wednesday, March 3, 2010, 8:18 PM
Hi Andrea,
Jackson lists a couple of FITC-conjugated donkey anti-rat secondaries:
712-095-150 - Whole Donkey Anti-Rat IgG (H+L).
712-096-150
- F(ab')2
. I
would give you the catalog number except I am not at my computer.
Andrea T. Hooper
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. I
would give you the catalog number except I am not at my computer.
Andrea T. Hooper
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. anonwu...@gmail.com
Subject: Re: [Histonet] Snap Freezing Tissue
To: Andrea T. Hooper andreahoo...@rocketmail.com
Cc: histonet@lists.utsouthwestern.edu, Laurie Colbert
laurie.colb...@huntingtonhospital.com, Steve Pike ste...@mikronet.com
Date: Wednesday, February 24, 2010, 10:31 PM
I learned
the autofluorescence from
RBCs. I am afraid of false positives.
--- On Tue, 2/23/10, Adam . anonwu...@gmail.com wrote:
From: Adam . anonwu...@gmail.com
Subject: Re: [Histonet] Double labeling with antibodies that need different
fixatives
To: Andrea T. Hooper andreahoo...@rocketmail.com
Cc: Phebe Verbrugghe
What types of mouse tissue are you working with? Are they fresh frozen with
post-fixation or fixed frozen? Do you block biotin? Do you have an isotype
control and a no primary control? I am thinking biotin's the likely culprit as
the kit doesn't come with the very necessary biotin blocking
Not sure if you were asking me or not and it's a broad answer but VE-cadherin
is a good one - the best overall marker of endothelium. vWF is another one that
works well for most tumors. If staining mouse organs, if you let me know what
tissue you are staining I can recommend the best specific
.
If snap freezing tissue for RNA/protein extraction I use the classic isopentane
cooling method.
Andrea T. Hooper
--- On Wed, 2/24/10, Laurie Colbert laurie.colb...@huntingtonhospital.com
wrote:
From: Laurie Colbert laurie.colb...@huntingtonhospital.com
Subject: [Histonet] Snap Freezing
Hi Phebe,
In my experience that CD31 clone you refer to doesn't work well mainly b/c of
paraffin embedding - in combination with PFA/formalin fixation. On
PFA/formalin fixed frozen samples it works just fine with a trypsin antigen
retrieval step. I find it's the paraffin that is the real
For many years doing IHC on mouse bones, I have used 10% EDTA, just pHed up to
where it goes into solution, with great results for frozen section
immunofluorescence. It takes 5 days. For FFPE, we use Richard Allan Decal
solution which is HCl and EDTA. Also works great and only takes 1 hour for
After cytospinning I always dry my slides thoroughly before moving on to a IHC
or IF step. I also use Superfrost Plus slides or silane coated slides to
increase adherance. I would do that then try various fixatives.
You can make a cell pellet as you suggest. Decide ahead of time if you want to
Hi Julia,
I am sorry it's taken me months to reply to this email and hopefully it's not
too late. I am only catching up on Histonet messages recently.
I stain for VE-cadherin, CD144 in murine bone routinely. RD Systems goat
anti-mouse VE-cadherin works very well in paraffin and frozen murine
Although what you say is true, it is worth noting that there are some
antigens/antibodies more amenable to acid decal. I have experienced this myself
several times recently and been burned by thinking EDTA will always give better
IHC results. At least for murine bone marrow vasculature
Couldn't agree with Gayle more!
Glucose oxidase is the best protocol for blocking endogenous peroxidase but
isn't entirely necessary unless you are working with hematopoietic tissue or
tissue that has a large leukocyte infiltrate. For muscle standard 0.3%
H202 should be fine. Of course if you
Your protocol is fine. It's precisely my protocol.
What do you get when you do the no primary control? Although this control isn't
sufficient for publication it is necessary for troubleshooting. In fact
performing the following controls together can tell you exactly where the
problem lies:
Hi everyone! It's good to be back ... I have resubscribed under a different
email as I switched place of employment.
I am just running a poll as to what your favorite anti-VEGFR2 antibody is for
immunostaining human tissue or cells. Frozen or paraffin, cells or tissues
... it's all good info
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