About 3 years ago I ask a vendor who was selling polymers for animal tissue if
I could get it with fluchromes. He told me it couldn't be done. I haven't seen
any on the market so I am guessing no one has figured out how to do it. It
would sure be nice for research work.
Donna Reynolds HT(ASCP)
We are a research lab and still do all our staining by hand. Just prefer
it that way.
Our carpenter shop makes ours. We determine size by how many slides we want
to stain at one time. We have chambers that will hold 5, 30, 40, 60 slides.
We make the Plexiglas box out of black Plexiglas
I have an investigator wanting to make microarrays from some frozen OCT blocks.
I seem to remember reading a way to do this. Can someone please point me in the
right direction?
Thanks
Donna Reynolds, HT (ASCP) Chief IHC Core Lab
Dept. Cancer Biology,
Univ. Tex. M.D. Anderson Cancer Center
I recently tested a mCD34 from GeneTex cat GTX28158 that gave very nice
labeling of mouse endothelial cells on FFPE. Used EDTA pH 8 antigen retrieval
and no amplification.
For those of you using the Dianovo mCD31 antibody what are you using for
antigen retrieval? And are you using
What are you using for Endogenous blocking? If you are using methanol with H2O2
that may be your problem. Many CD antibodies are very sensitive to the
methanol. Try PBS or whatever buffer you are using to dilute the H202. I would
also try going straight form the -20 into cold acetone without
I am not sure what all is in the kits. We rarely buy kits, they are expensive
and often as not you can get better reagents by buying them individually.
For our HRP-IHC we buy HRP conjugated antibodies from Jackson Immuno. They have
a large selection, reasonably priced, and work great.
For AEC
Is there a block alignment tool that will work in a cryostat? I work in
research and frequently have to go back and recut small pieces of tissue. My
Leica allows me to adjust the head but it is very difficult to get the block
lined up on the same axis as before.
Donna Reynolds Core IHC Lab
I cut mouse brains frozen in OCT with very little problem. I bring the
Cyrostat temp up to between -15 to -13. Pick them up with a warm slide and
seldom have a wrinkle. Brains are unfixed placed in OCT and frozen with liquid
nitrogen.
Donna Reynolds, Chief Histology Lab (IHC)
U.T, M.D.
I am not familiar with what IHC world has on this but we have used this
technique for a number of years. The mouse fragment we use is from Jackson cat
#115-007-003. We dilute it 1:10 in our protein blocking solution. It works
better with DAB than with Fluorescence. We have also found that to
For MMP2 we use Rabbit anti-MMP2 - Abcam ab79781 with pepsin digestion 1:400
with Jackson goat anti-Rabbit HRP secondary.
Message: 2
Date: Fri, 16 Mar 2012 17:40:28 +
From: Chiriboga, Luis luis.chirib...@nyumc.org
Subject: [Histonet] MMP2
To: 'histonet@lists.utsouthwestern.edu'
I don't know about implants but we were having a terrible time with bone
marrows coming off. Usually they came off during the HEIR using the steamer.
At that point we were placing our slides lying flat in a 55-60º oven overnight
which usually helps tremendously with difficult tissue. This was
We have a system we make that uses Plexiglas boxes to create the humid
environment and a special holder inside that holds 1-10 slides that can be
picked up and washed all at once.
We have our new baby chamber that only holds 5 slides for the minnie jobs. Our
chambers will hold 3 holders =30
We do this frequently.
Fix slides in Methanol (45ml) + formaldehyde (5ml) for 10 minutes.
Wash several changes of tap water
Stain with Harris Hematoxylin for 3 minutes
Proceed as you would for paraffin sections
Get great staining.
I am sure if you are using a different Hematoxylin you could
I don't work with these antibodies but when we were doing bone marrow bx
pressure cooker, high pH retrieval we lost a lot of tissue. This was what we
had previously work out for our antibody. We replaced the pressure cooker with
a 70ºC water bath, kept the high pH retrieval and cooked for 2
I just learned the company making the Brigati Stable DAB is no longer making
it. Does anyone know if someone else has been able to pick up this formulation
and produce it or are planning to make it. We have been using this for over 20
years and I really hate to change. What DAB are others using
We have a motorized and rarely use the motorized function, Maybe 5 time in the
last 5 years. It is faster to cut manually.
Donna Reynolds, chief Histology research lab
U.T. M.D. Anderson Cancer Center, Houston, TX
713-792-8106
___
Histonet mailing
I is my understanding that the Mach 3 is for use on human clinical samples and
will not work in mouse tissue. Maybe BioCare technical person didn't realize
you were using human cell in the mouse. BioCare has a great polymer for rabbit
antibodies on rodent tissue in their ProMark series. Rabbit
Has anyone ever experienced OCT blocks that cut like they are soft. I have
turned the cyrostat down 10 degrees lower than usual and they are still soft.
Really weird part is that I have 20 blocks in this group and some are soft and
others are just fine. Tried changing angle, new blade nothing
We had a similar problem a few years back. We had used an antibody at 1:500
dilution for years. Got in a new bottle and no labeling. Called the
manufacture and they assured me it was the same concentration. Finally went
to 1:50 dilution and the staining was like our old stain. Over the last
We had the same problem with our Leica 3050S. It happened occasionally then
became more and more frequent. We went through a lot of the same hoops you have
gone through. When it shut down one day while I was cutting I knew it had to be
a machine malfunction. Leica replaced something that was
I have tried several brands all have the same problem some worse than others.
But even worse than the coverslips are the slides. I cut a lot of frozen
sections on charged slides for fluorescent labeling the slides are dirty and
the dirt gets trapped under the tissue as well as around it. The
From: Reynolds,Donna M
Sent: Friday, September 18, 2009 8:01 AM
To: 'ih...@googlegroups.com'
Subject: RE: Use of immuno cal or other gentle formic acid decal solution for
immuno specimens
I have attached a decal procedure we have used for a long time. We use it for
mouse bones mostly. But we
We have used mouse anti-Brdu from Becton Dickinson for years. It is still
available from BD BioSciences Cat. 347580. I know it is made in mouse but we
used an IgG1 secondary and always had good results. There may be some
cytoplasmic label, probably background, but you are interested in the
Research Genetics has merged a couple of time the last few years. They are now
Open Biosystems in Huntsville, Al pho 888 412 2225. They do not list the
Research Genetics on the Web but last I knew products were still available. You
can call them to get current catalog number and prices.
Donna
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