[Histonet] Cryostat disinfection
In my previous email regarding weekly cryostat disinfection-- I meant to say that we clean with 70% alcohol, then bring to room temp, disinfect with Cavicide, wipe down with abs alcohol, bring back down to -22C. We also use 70% alcohol for our routine daily cleaning. Cindy Baranowski Emory Healthcare Sent from my iPad ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Biocare Decloaker Nx Gen
We recently bought this equipment and it works well. We are happy with it. -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Blazek, Linda Sent: Thursday, March 28, 2013 1:52 PM To: Swartwood, Steven J; histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: Biocare Decloaker Nx Gen We have used the Decloaking Chamber for several years now and have had excellent results. The nicest thing is that there is consistency and reliability. Each run will be the same as the one previous and having the ability to use the USB stick for documentation of time, pressure and temperature makes record keeping really easy. We have had no complaints. The only con I can come up with is that it comes in a big box that is hard to store if you have to send it back for some reason. Linda Blazek HT (ASCP) Manager/Supervisor GI Pathology of Dayton Digestive Specialists, Inc Phone: (937) 396-2623 Email: lbla...@digestivespecialists.com -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Swartwood, Steven J Sent: Thursday, March 28, 2013 1:40 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Biocare Decloaker Nx Gen Has anyone ever used the Biocare Decloaking Chamber Nx Gen for epitope unmasking (heat retrieval)? Also does anyone have any personal pros/cons with this instrument? We are thinking about getting one because of inconsistencies with our current method using a vegetable steamer. We use many different tissue types both animal and human. Also we use many different antibodies. Currently with the vegetable steamer we've used Citrate, EDTA, and Tris-EDTA combinations. We've used times from 30-60 minutes, and even tried 30 minutes x 2 or 40 minutes x 2, but we still see some inconsistent staining. To make sure it wasn't due to fixation vimentin staining was performed and shows even staining. It's just certain antibodies that are giving us trouble even from multiple different companies. Any advice would be greatly appreciated and very beneficial to our work pertaining to the instrument. Thanks again everyone have a great end of the week, Steven Swartwood HT(ASCP) steven.swartw...@cshs.org IMPORTANT WARNING: This message is intended for the use of the person or entity to which it is addressed and may contain information that is privileged and confidential, the disclosure of which is governed by applicable law. If the reader of this message is not the intended recipient, or the employee or agent responsible for delivering it to the intended recipient, you are hereby notified that any dissemination, distribution or copying of this information is STRICTLY PROHIBITED. If you have received this message in error, please notify us immediately by calling (310) 423-6428 and destroy the related message. Thank You for your cooperation. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] EDTA decal
Good idea. I have to soak the edta decaled tissues in an alkaline solution
inorder to restore enzyme histochemical staining for TRAP, might be the same
issue for some IHC.
Patsy Ruegg, HT(ASCP)QIHC
Ruegg IHC Consulting
40864 E. Arkansas Ave
Bennett, CO 80102
H 303-644-4538
C 720-281-5406
prueg...@hotmail.com
rueggihcconsultin...@outlook.com
This email is confidential and intended solely for the use of the Person(s)
('the intended recipient') to whom it was addressed. Any views or opinions
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-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Teri Johnson
Sent: Friday, August 16, 2013 11:54 AM
To: 'cfors...@umn.edu'
Cc: histonet@lists.utsouthwestern.edu
Subject: [Histonet] EDTA decal
Hi Colleen,
I would say it's unusual, but not completely impossible that EDTA has
interfered with your IHC. We had that problem with demonstrating
B-galactosidase in mouse bones. If we decalcified it in EDTA after whole
mount staining in X-gal, the blue staining was removed. But if we
decalcified it in formic acid, the stain was retained.
I think a few years ago, Biogenex had a room temperature antigen retrieval
solution for acid decalcified bone (not the same as your situation, I know).
But I think it was largely an alkaline solution you let the slides sit for
maybe 30 minutes prior to staining. Might be worthwhile to try a different
retrieval method for these and see what happens.
Otherwise, are you sure you are using a clone that reacts in mouse tissue?
We use Rabbit monoclonal SP6.
Teri Johnson
Manager, Histology
GNF - San Diego, CA
858-332-4752
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RE: [Histonet] IHC after EDTA decalcifying
Colleen,
The antigen expression you are getting in the human tonsil may not be
expressed in the mouse femur, plus u have to make sure the ab u are using
cross reacts with mouse tissue, if it is anti human it may or may not react
in mouse, the ab data sheet should indicate species reactivity. The edta
should not be the problem here.
Patsy
Patsy Ruegg, HT(ASCP)QIHC
Ruegg IHC Consulting
40864 E. Arkansas Ave
Bennett, CO 80102
H 303-644-4538
C 720-281-5406
prueg...@hotmail.com
rueggihcconsultin...@outlook.com
This email is confidential and intended solely for the use of the Person(s)
('the intended recipient') to whom it was addressed. Any views or opinions
presented are solely those of the author. It may contain information that is
privileged & confidential within the meaning of applicable law. Accordingly
any dissemination, distribution, copying, or other use of this message, or
any of its contents, by any person other than the intended recipient may
constitute a breach of civil or criminal law and is strictly prohibited. If
you are NOT the intended recipient please contact the sender and dispose of
this e-mail as soon as possible.
-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Colleen
Forster
Sent: Thursday, August 15, 2013 5:59 PM
To: Histonet
Subject: [Histonet] IHC after EDTA decalcifying
Hello fellow histonetters,
Can someone who has worked with decalcifyied bone tell me if EDTA interferes
with IHC staining? I was under the impression it did not but cannot get
staining in mouse femurs that I have decaled in EDTA. I have used both the
steamer and the decloaker for retrieval and in the human tonsil the stain is
great.
Any suggestions..
Thanks in advance!
Colleen Forster
U of MN
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[Histonet] Cryostat cleaning
The only method acceptable to CAP is bringing the cryostat to room temp and disinfecting weekly UV light will not be an acceptable alternative by CAP. We clean out all tissue debris, wipe down with abs alcohol, bring cryostat to room temp, use Cavicide to disinfect, wipe down again with abs alcohol. We use 70 % alcohol for daily cleaning. Cindy Baranowski Emory Healthare Atlanta, GA Sent from my iPad ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] edge artefact on core needle biopsy
Hi all! We had a discussion on the cause of IHC positivity/false-positivity on the edges of core needle biopsies. (eg. breast, ER) One said, it is due to overfixation with formaldehyde-concentrations above 5%. (=false pos.) My opinion is, that's it's rather an effect of underfixation in the center in those cases. (= real pos.) In my lab, we rarely see this margin-effect and the biopsies are fixed for 6 hours at least. I searched in Pubmed for publications on this assay, but without success. Can anyone help me out with any literature? Or personal experience? Thanks in advance Gudrun Lang Ltd. BMA Histolab Linz, Austria ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
AW: [Histonet] post-fixation for Mallory Trichrome
Hi, I think staining would be improved, when you use Bouin as postfixative ( 1h hour, 60°C). Try to invite a further step between Fuchsin and Anilinblue. When you incubate the slides in 1% phosphomolybdic acid for 5-10 min, you can see decolorization of cardilage. This improves later on anilin-binding. You may control the differentiation-step under microscope. Anilinblue can be reduced to 5 min afterwards. You may also consider, that there's a real difference in cardilage-architecture of the early and late samples, that takes influence on staining-abilities. Gudrun Lang -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Rui TAHARA Gesendet: Freitag, 16. August 2013 23:33 An: histonet@lists.utsouthwestern.edu Betreff: [Histonet] post-fixation for Mallory Trichrome Hello, There seems to be a lot of suggestions for Mallory Trichrome staining involved in Acid fusin, Aniline blue, and orange G. My sample of avian embryos were fixed with Formalin based fixative (4% paraformaldehyde in phosphate-buffered saline and 1% glutaraldehyde) overnight, in case of late embryos bones were decalcified, and followed standard alcohol series, paraffin embedded, and 10 micron sectioned. The slides were dehydrated, stained with 0.5 % Acid fushin for 10min, then without wash, 0.5% Aniline blue and 2 % orange G in 1% phosphomolybdic acid solution for 20 min, then undergone ethanol series, cleared and mounted. Now I know that formalin fixed sample do not present optimized trichrome colors based on your websites and references. Yet the stained sections of late stage embryo do still show near optimized colors whereas those of early stage embryo show almost no blue or very dark blue, almost gray color for cartilages and most of morphologies as purple-redish colors. Since I tested staining on late embryos first I thought staining would work on early embryos. Does anyone provide me explanation why the staining mostly shows red-ish color and not optimized color for cartilage in early embryos? Is that because of the formalin-fixed embryos although late stage embryos fixed with formalin still show the blue for cartilage? Another question is related to fixative. I prefer not to use bousin fixative due to picric acid and it seems that Citrate buffer or Grams iodine solution can be substituted to bousin post-fixative. Have anyone tried these solution for Mallory Trichrome? Do those methods show near optimized color for Mallory trichrome? Any suggestion is appreciated. Thank you, Rui TAHARA PhD candidate McGill University, Biology Department ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
AW: [Histonet] EDTA decal
We experienced different IHC staining qualities on bone marrow, that was decalcified with EDTA or formic acid. I remember, our CD38 antibody was always very weak after EDTA and made no problems after formic acid. I think it is very difficult to make an over-all-statement for all antigens/all antibodies. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Teri Johnson Gesendet: Freitag, 16. August 2013 19:54 An: 'cfors...@umn.edu' Cc: histonet@lists.utsouthwestern.edu Betreff: [Histonet] EDTA decal Hi Colleen, I would say it's unusual, but not completely impossible that EDTA has interfered with your IHC. We had that problem with demonstrating B-galactosidase in mouse bones. If we decalcified it in EDTA after whole mount staining in X-gal, the blue staining was removed. But if we decalcified it in formic acid, the stain was retained. I think a few years ago, Biogenex had a room temperature antigen retrieval solution for acid decalcified bone (not the same as your situation, I know). But I think it was largely an alkaline solution you let the slides sit for maybe 30 minutes prior to staining. Might be worthwhile to try a different retrieval method for these and see what happens. Otherwise, are you sure you are using a clone that reacts in mouse tissue? We use Rabbit monoclonal SP6. Teri Johnson Manager, Histology GNF - San Diego, CA 858-332-4752 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
