I have cleaned my disposable Miles Blades by holding blade with disposable
forceps and dipping in Xylene first---wipe with kimwipe; then absolute
alcohol---kimwipe----this removes any oils if I am cutting to extract RNA
either paraffin or frozens---hi Freida!!

                                With warmest regards,
				Louise "Beezie" Burrell
				Chief Technologist-Cancer Center
				Tissue Procurement Core Facility
				Campus Box #8056
				Washington University School of Medicine
				St. Louis, MO.  63110
				Ph:  314-454-7615
				Fax: 314-454-5525

On Wed, 7 Apr 1999, HistoNet Server wrote:

> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 00:08:02 -0600
> From: "M. Brown" <marianb@u.washington.edu>
> Subject: Re: Re; endogenous peroxidase blocking
> 
> I have already written the just about the same reply as far as blocking
> after the 2nd antibody(Link). I did however dilute the H2O2 with Methanol
> only for bone marrow smears and biopsy cores.
> Marianne
> 
> 
> 
> 
> On Sat, 27 Mar 1999, bbracing wrote:
> 
> > Cynthia,
> > Back in the late 80's early 90's when IHC was still in its infancy and about
> the only reliable IHC method used the P.A.P. technology, I did quite an
> extensive comparison study on a large number of tumors using many different
> antibodies, and in many cases it was quite easy to demonstrate that blocking
> with H2O2 before the primary antibody was applied,  had quite a negative
> affect on many tumors.  I was also able to demonstrate that H2O2 blocking
> after the primary antibody, but before the linking antibody also had quite an
> negative affect on the detection of many tumor antigens. I was not able to
> demonstrate any negative effects if the blocking step was placed between the
> link and lable reagents, and so for the past ten or so years this is where we
> do our endogenous peroxidase blocking.  With the advent of the streptavidin
> biotin technology, I found that with the increased sensitivity this problem
> was slightly reduced, but I still kept the blocking step between the link and
> lable steps. For the last couple of years I have been using the Labelled
> Polymer (Dako) technology (which is a one step technology that solved several
> problems we were having with the link lable methods) but I still found it
> beneficial to block after the primary, but before the labelled polymer. 
> However, I have not done any comparison studies on tumors which have had heat
> induced antigen retrieval techiques applied  to them, to see if the placement
> of the H2O2 blocking step, has any effects on the heat retrieved antigens. 
> This might be an interesting project?
> > Anyway, one other point.  I have never been given a satifactory answere as
> to why the 30% H2O2 in many labs is diluted with absolute, or in some cases
> 70% alcohol. The naturally occurring  peroxidase enzyme works in an aqueous
> environment so why not dilute the H2O2 with distilled water, or saline?  I use
> the former, and to the best of my knowledge it works just fine and is a lot
> less hassle, as a 25 ml bottle lasts a week, before I have to make up fresh.
> > Kerry Beebe
> > Kelowna Gen Hospital
> > B.C. Canada
> > 
> > 
> > 
> > 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 04:15:53 -0600
> From: OhioHisto@aol.com
> Subject: Ohio Conference
> 
> Dear Histonetters, 
> 
> The Event:
> You are invited to attend the Histology Society of Ohio 25th Annual 
> Conference which will be held on June 24-27 at Mohican Resort and Conference 
> Center.  Please come and visit our Web site at 
> http://www.hsoweb.org/conf99/index.htm for more information and to register 
> to  attend or request a program online.  What are we going to talk about? 
> Naturally Histology!  This conference will not only focus on histology, it 
> will cover all aspects of anatomic pathology (both veterinary and human) and 
> some personal growth and wellness issues. The conference committee has come 
> up with the most innovative topics presented by world renowned people who are 
> leaders in their field of interest.  Something that you don't want to miss in 
> this beautiful relaxing atmosphere.
> 
> Location:
> The Mohican Resort, a national gold medal award winner for state parks and 
> recreation excellence, is located just 20 miles southeast of Mansfield.  A 
> one hour drive north of Columbus, Ohio and two hours drive south of 
> Cleveland, Ohio.  Whether a family vacation, weekend getaway, one-day 
> workshop, or at a 3-day conference, the Mohican staff will be pleased to 
> provide you with a pleasant, successful and meaningful stay.  The discounted 
> Resort rate is $120 per room plus tax. To make your Resort reservations, the 
> deadline is April 25, 1999.  Please call Mohican Reservations at 
> 1-800-282-7275.
> 
> If you have any questions, please feel free to contact us.  We hope to see 
> you there. 
> 
> You may contact: 
> 
> Elizabeth Wenig 
> E-mail: conf99@hsoweb.org 
> Ph: 614.566.9177 
> Fax: 614.566.8862 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 04:25:41 -0600
> From: "Marshall, Sharon, Mrs" <marshall@anat.uct.ac.za>
> Subject: mallory trichrome method
> 
> Hi  histonetters,
> 
> I wonder if anybody has the method for the mallory trichrome stain  
> which uses acid fuchsin(red cells), phosphotungstic acid and aniline 
> blue-orange G(muscle etc.). I have lost my method and I am not sure 
> of the exact times of the tissue sections in each. 
> 
> Thanks
> Sharon Marshall
> e-mail: marshall@anat.uct.ac.za
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 04:35:17 -0600
> From: "Saby, Joseph" <Joseph.Saby@wl.com>
> Subject: Position Available:  Corrected Listing
> 
> Please note that the ideal candidate should have 2 years of professional
> experience.
> 
> 
> 
> Parke-Davis Pharmaceutical Research, a division of Warner-Lambert Company
> located in Ann Arbor, MI has an immediate opening for an Assistant
> Scientist/Histotechnologist.  In accordance with established procedures you
> will be responsible for procuring and preparing animal tissue sections using
> routine and special techniques for examination by light microscopy.
> 
> The ideal candidate has a minimum of 2 years of professional experience. A
> BS degree and HT or HTL ASCP certification are preferred.
> 
> Qualified candidates mail, fax or e-mail your resume, indicating Job Code:
> CJMJJ to: Parke-Davis, 2800 Plymouth Road, Ann Arbor, MI 48105.  FAX (734)
> 622-7617.  E-mail: Human Resources.resume@wl.com.
> 
> 
> Joe Saby, BA HT
> Parke-Davis, Ann Arbor, MI
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 04:35:50 -0600
> From: afbrand@liverpool.ac.uk (A. F. Brandwood)
> Subject: DiaChem/Diagnostic Developments
> 
> Dear Eva,
> 
> DiaChem's address is:-
> 
> DiaChem International Ltd,
> Unit 5,
> Gardiners Place,
> West Gillibrands,
> Skelmersdale,
> Lancashire,
> WN8 9SP.
> 
> Tel No 01695 555581
> Fax No 01695 555518
> E-Mail:- diachem@cybase.co.uk
> 
> Best Wishes,
> 
> Tony Brandwood,
> Dept. of Vet. Pathology,
> University of Liverpool,
> P.O. Box 147,
> Liverpool,
> L69 3BX,
> U.K.
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 06:15:48 -0600
> From: "Tim Morken" <timcdc@hotmail.com>
> Subject: Re: endogenous peroxidase,
> 
> Bruce,
> 
> You are right in thinking that you can probably eliminate the h2o2 in 
> brain tissue IHC. It is mostly blood cells that cause the problem. In 
> perfused tissue it may be even less of a problem due to lack of leaching 
> that occurs between time of cell death and fixation. 
> 
> In fact, in any preparation in which you feel the h202 may cause 
> problems, you can put up with endogenous peroxidase staining after 
> determining what is endogenous and then ignoring it when evaluating the 
> slides.
> 
> Some even use the endogenous staining to their advantage as proof the 
> DAB (or other chromogen)is working correctly.
> 
> Tim Morken, B.A., EMT(MSA), HTL(ASCP) 
> Infectious Disease Pathology
> Centers for Disease Control
> MS-G32
> 1600 Clifton Rd.
> Atlanta, GA 30333
> USA
> 
> email: tim9@cdc.gov
>        timcdc@hotmail.com
> 
> FAX:  (404)639-3043
> 
> 
> 
> 
> 
> 
> 
> - ----Original Message Follows----
> From: Bruce A Rasmussen <brasmuss@osf1.gmu.edu>
> To: HistoNet@Pathology.swmed.edu
> Subject: endogenous peroxidase,
> Date: Mon, 05 Apr 1999 18:02:34 -0400 (EDT)
> 
> 
> Is background staining caused by endogenous peroxidase mainly a blood
> confound? In other words with well perfused brain tissue is H202
> unnecessary or not worth the tissue damage and loss of immunoreactivity?
> Many thanks,
> Bruce
> 
> 
> 
> - --------------------------------------
> Bruce Rasmussen
> Predoc Fellow in Experimental Neuropsychology
> The Krasnow Institute for Advanced Study
> George Mason University
> Mail Stop 2A1
> Fairfax, VA 22030-4444
> Office:703-993-4358
> Lab:703-993-4369
> Home:703-765-4570
> Fax:703-993-4325
> - ---------------------------------------
> 
> 
> 
> 
> 
> 
> Get Your Private, Free Email at http://www.hotmail.com
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 08:00:46 -0600
> From: afbrand@liverpool.ac.uk (A. F. Brandwood)
> Subject: Mallory trichrome
> 
> Dear Sharon,
> 
> The technique can be found in 'Handbook of histopathological and
> histochemical techniques by C.F.A. Culling, third edition, published by
> Butterworths, page 415.
> If you have any difficulty obtaining a copy of this book please e-mail me.
> 
> Best wishes,
> 
> Tony Brandwood,
> Dept. of Vet. Pathology,
> University of Liverpool,
> P.O. Box 147,
> Liverpool,
> L69 3BX,
> U.K.
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 09:01:04 -0600
> From: Bruce A Rasmussen <brasmuss@osf1.gmu.edu>
> Subject: Thanks re:endogenous peroxidase, a serum blocking question
> 
> My thanks to all who wrote back with detailed responses to my endogenous
> peroxidase question. If I could trouble folk with just one more question. 
> I have a protocol that calls for a 5% serum block (30 min to the secondary
> animal) before and primary and before the secondary. There is much debate
> about whether the second block is necessay, if it will reduce the signal,
> or if rinsing after the serum blocks is good or bad. Some folk also
> suggest that a couple of percent BSA is a good idea.
> Thanks again,
> Bruce 
> 
> 
> 
> - --------------------------------------
> Bruce Rasmussen
> Predoc Fellow in Experimental Neuropsychology
> The Krasnow Institute for Advanced Study
> George Mason University
> Mail Stop 2A1
> Fairfax, VA 22030-4444
> Office:703-993-4358
> Lab:703-993-4369
> Home:703-765-4570
> Fax:703-993-4325
> - ---------------------------------------
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 09:13:02 -0600
> From: Kathy Wortham <kjwortham@yahoo.com>
> Subject: Re: sticky slides
> 
> Procedure for silane coated slides.
> 
> 1-Place slides in staining racks
> 2-Place slides in acetone 2 minutes
> 3-Place slides in 2% silane solution 2 minutes
>   (5ml silane to 250 ml acetone)
> 4-Distilled water 5 dips
> 5-Distilled water 5 dips
> 6-Distilled water 5 dips
> 7-Allow slides to air dry overnight
> 8-Store in slide boxes
> 
> I order the Silane from sigma. (3-aminopropyl-triethoxysilane 
> Cat#A-3648 
> 
> I have tried commercially available silane and plus slides but they do
> not work as well foe me and they are A LOT more expensive.
> 
> Kathy Wortham
> USDA, Food Safety Inspection Service
> 950 College Station Road
> Athens, Ga 30605
> 706-546-3556
> 
> 
> - --- "Masayuki Miyagishima, MD" <mmiyagis+@pitt.edu> wrote:
> > Could you let me know how to make silane coated
> > slide?  Are there any
> > commercially availabld silane slide?  Is it better
> > than superfrost plus
> > slides?
> > 
> >
> > Masayuki Miyagishima, MD
> > University of Pittsburgh, 
> > Department of Surgery
> > 412-647-2345, and ask the hospital operator to page
> > 3228, please.
> > 
> 
> ===
> 
> _________________________________________________________
> Do You Yahoo!?
> Get your free @yahoo.com address at http://mail.yahoo.com
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 09:45:37 -0600
> From: "Tim Morken" <timcdc@hotmail.com>
> Subject: Re: Thanks re:endogenous peroxidase, a serum blocking question
> 
> Bruce,
> 
> We use a 20% sheep serum (in tris-saline-tween20 buffer) before the 
> primary. We dilute the primary in the same serum-buffer, so it 'keeps on 
> blocking.' We don't rinse the blocking serum before putting on the 
> primary, only an air-blow (using the DAKO auto-stainer). We then rinse 
> the with the tris-saline buffer without the serum.
> 
> If you have a lot of background problems then a second serum blocker may 
> be necessary before applying the secondary antibody. In general, I would 
> rather keep the procedure simple and use as few steps as possible.
> 
> Tim Morken, B.A., EMT(MSA), HTL(ASCP) 
> Infectious Disease Pathology
> Centers for Disease Control
> MS-G32
> 1600 Clifton Rd.
> Atlanta, GA 30333
> USA
> 
> email: tim9@cdc.gov
>        timcdc@hotmail.com
> 
> FAX:  (404)639-3043
> 
> 
> 
> 
> - ----Original Message Follows----
> From: Bruce A Rasmussen <brasmuss@osf1.gmu.edu>
> To: HistoNet@Pathology.swmed.edu
> Subject: Thanks re:endogenous peroxidase, a serum blocking question
> Date: Tue, 06 Apr 1999 10:44:30 -0400 (EDT)
> 
> My thanks to all who wrote back with detailed responses to my endogenous
> peroxidase question. If I could trouble folk with just one more 
> question. 
> I have a protocol that calls for a 5% serum block (30 min to the 
> secondary
> animal) before and primary and before the secondary. There is much 
> debate
> about whether the second block is necessay, if it will reduce the 
> signal,
> or if rinsing after the serum blocks is good or bad. Some folk also
> suggest that a couple of percent BSA is a good idea.
> Thanks again,
> Bruce 
> 
> 
> 
> - --------------------------------------
> Bruce Rasmussen
> Predoc Fellow in Experimental Neuropsychology
> The Krasnow Institute for Advanced Study
> George Mason University
> Mail Stop 2A1
> Fairfax, VA 22030-4444
> Office:703-993-4358
> Lab:703-993-4369
> Home:703-765-4570
> Fax:703-993-4325
> - ---------------------------------------
> 
> 
> 
> 
> 
> Get Your Private, Free Email at http://www.hotmail.com
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 10:30:36 -0600
> From: Gayle Callis <uvsgc@msu.oscs.montana.edu>
> Subject: F4/80 for mouse macrophages.
> 
> We do it routinely, but only on frozen sections.  
> 
> NBF/paraffin sections generally don't work well with mouse cell surface
> markers.  We also grow up our own antibody and stain with the supernate. 
> It is available from SeroTec.  
> 
> It may work better with Carnoys fixation, instead of NBF.  We avoid 
> formalin for all our mouse markers with far better results.  F4/80 is
> also known to work with a FORMALIN FREE fixative containing zinc chloride,
> zinc acetate, calcium acetate in TRIS buffer then paraffin sectioning.
> 
> If you are interested in the frozen section protocol, will be glad to share
> ours.
> 
> Gayle Callis
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 11:15:47 -0600
> From: bwright@gene.COM
> Subject: Re: F4/80 murine mac marker
> 
> Brett,
> 
> We successfully stain with F4/80 (Serotec #MCAP497) on paraffin embedded
> mouse tissues.  We use spleen as a positive control.  0.4% pepsin at 37C
> for 10 minutes for antigen retrieval and an overnight  at 4C incubation
> in a 10ug/ml dilution of F4/80.
> 
> If you want the procedure just let me know.
> 
> Thanks
> Barb Wright
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 11:16:10 -0600
> From: FreidaC@aol.com
> Subject: Cleaning of disposable blades
> 
> I am very curious as to how many of you clean your disposable blades before 
> use.  If you do clean, what do you use?
> 
> Thanks for answering my curiosity.
> 
> Freida Carson
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 11:45:33 -0600
> From: "M. Brown" <marianb@u.washington.edu>
> Subject: Re: Cleaning of disposable blades
> 
> I usually just run the edge of blade across one side of a paraffin block,
> or run a piece of folded tissue paper across edge. When I first started
> using disp.blades, this is what I was told to do as sometimes there are
> very small particles of metal. Depending on the brand, I do not have to do
> this all the time however.
> Marianne
> 
> 
> 
> 
> On Tue, 6 Apr 1999 FreidaC@aol.com wrote:
> 
> > I am very curious as to how many of you clean your disposable blades before 
> > use.  If you do clean, what do you use?
> > 
> > Thanks for answering my curiosity.
> > 
> > Freida Carson
> > 
> > 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 11:56:12 -0600
> From: amdj@duke.edu
> Subject: Re: Cleaning of disposable blades
> 
> I find that the safest method is to run the blade through a clean chunk 
> of paraffin or a blank block. I have used gauze, terriwipes, antistatic 
> pads and a heavy cotton towel to wipe the blade. All are more damaging 
> to the blade and my fingertips.  Bert Dotson
> 
> On Tue, 06 Apr 1999 12:57:20 -0400 (EDT) FreidaC@aol.com wrote:
> 
> > I am very curious as to how many of you clean your disposable blades before 
> > use.  If you do clean, what do you use?
> > 
> > Thanks for answering my curiosity.
> > 
> > Freida Carson
> > 
> 
> - ----------------------
> Bert Dotson
> amdj@duke.edu
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 12:08:12 -0600
> From: "Williams, Matthew J [PRI]" <MWillia7@prius.jnj.com>
> Subject: exsanguination
> 
> 'Net folk-
> 
> I am looking for any documentation that explains the reasons for and
> supports the process of exsanguination of laboratory animals before a
> necropsy. A discussion has arisen over this subject, and the possibilty that
> it is an inhumane and unduly painful method (even though the animals are
> anesthetized with sodium thiopental). If you care to get back to me directly
> my address is Mwillia7@prius.jnj.com.
> 
> Thanks!
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 12:19:19 -0600
> From: "Goodwin, Diana" <DGoodwin@CHSNJ.org>
> Subject: RE: Progesterone clone 1A6
> 
> Joanne,
> 
> What dilution are you using?  I use the novacastra 1A6 @ 1:80 for 25 min
> @ room temp. with very good results.
> 
> 
> 
> 
> Diana Goodwin, HT
> Trenton, NJ
> 
> > ----------
> > From: 	Suder, Joanne[SMTP:JC_Suder@fccc.edu]
> > Sent: 	Monday, April 05, 1999 1:21 PM
> > To: 	'histonet@pathology.swmed.edu'
> > Subject: 	Progesterone clone 1A6
> > 
> > Histonetters,
> >   Having a background problem with PR purchased through Novacastra
> > clone
> > 1A6. Our routine has the slides in Ab overnight at 4o. We have
> > recently been
> > seeing background staining so we tried it a both rt and 4o. We're
> > still
> > getting the background staining. Any suggestions?? Please reply
> > JC_SUDER@FCCC.EDU
> > 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 12:31:29 -0600
> From: "Sarah Christo" <schristo@cvm.tamu.edu>
> Subject: Metallothionein
> 
> Dear Histonetters,
>    Does anyone know of histologic techniques for a heavy metal binding protein
> called metallothionein.  
> Thanks, Sarah
> 
> Sarah Christo, HT (ASCP)
> Texas A&M University
> College of Veterinary Medicine
> Dept. of Vet. Anatomy & Public Health
> College Station, TX  77868-4458
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 12:45:24 -0600
> From: larisonk@UONEURO.uoregon.edu
> Subject: Re: GFP/mounting media
> 
> Histonetters and Gayle,
> 
> Most antifade reagents quench fluorescence, while preventing further
> bleaching.  
> For example, I found that Vectashield quenches the flourescence of FITC by 
> about 50% (I used an image processor and a capillary tube full of reagents to 
> make these determinations), but is very good at stabilizing this reduced
> signal 
> over time.  That's the beauty of ProLong -- it really doesn't quench the 
> fluorescence of FITC much at all, while still providing good anti-fade 
> properties.  This is a product I developed when I worked at Molecular Probes, 
> so I'm familiar with it's attributes.  It's hard to predict how it would
> behave 
> with GFP, however.  I presume, the GFP protein directs electrons along a 
> certain pathway, much like rhodopsin or the phycobiliproteins.  The medium may
> 
> inhibit conformational changes in the protein required for directed electron 
> movement, or the antifade reagent itself may wreak havoc with the process.  In
> 
> general, my experience with anti-fades (and I've had plenty of experience) is 
> that they are extremely subject to environmental conditions.  For instance, an
> 
> antifade that works well with one antibody may not work well with another. 
> The 
> assumption is that there is some environmental factor at work.  For instance, 
> FITC fades extremely rapidly in some environments and is relatively stable in 
> others.  I assume the local redox environment is the the primary factor, as 
> fluorescence entails the excitation of electrons.
> 
> Karen Larison at University of Oregon
> 
> 
> Date:          Sat, 03 Apr 1999 17:23:12 -0700
> From:          Gayle Callis <uvsgc@msu.oscs.montana.edu>
> Subject:       GFP/mounting media
> To:            histonet@pathology.swmed.edu
> 
> We had the same problem with GFP and aqueous mounting media.  We ended up
> testing several aqueous media to see if fluorescence was reduced. Tested were
> PBS, PBS/glycerol 1:1, Aquamount, Gel mount, Crystal Mount (actually 
> recommended by Tech services at Clontech - GFP gurus) and Vectashield.
> 
> Results were the PBS (coverslip sealed with nail polish) maintained the
> flourescence best ( and we use GFP filters on the UV scope).  All others 
> showed a reduced fluorescence, very disappointing, especially  when
> Clontech said should work! The PBS/glycerol use was on sections of brain
> that were 300um thick, so the mounting media may not have penetrated this
> thick sections easily and the GFP fluorescence was maintained.  My sections
> were 5 um containing bacteria with GFP, then added problem of another 
> fluorescent marker on cells (IFA staining).  
> 
> 
> Molecular Probes sends out a tips on mounting media brochure with purchase of
> their Alexa 488 and 546 fluorochromes, that gives info on which media
> work best with the Alexa's, FITC and TRITC.  They include their Prolong,
> and antifade product.  Since we wanted to do an immunofluorescent stain
> on same tissue containing GFP, this info was important.
> We basically did not want to purchase their mounting
> media, pricey, particularly if it did not help maintain the GFP and have it
> sit around being a shelf ornament.
> 
> At this point, we live with the PBS mount, excellent for Alexa 488 and
> 546, take photos immediately on a computerized system.  Hopefully
> they have more info on their mounting media and GFP now, my inquiry did
> not get anywhere, since they are more involved with their fluorescent
> markers/dyes. If you try the Prolong,, I will be interested in your
> results.    
> 
> One sidelight, the Strepavidin Alexa conjugates are very nice, we like them
> better than TRITC, brighter, and works right into our immunostaining
> protocols with biotinylated primaries or secondaries.  High dilutions, better
> We have GFP in bacteria and did an immunostain on same section using 
> Alexa 546 (TRITC replacement) with super results
> 
> The GFP seems sensitive to some of the ingredients in mounting media,
> polyvinyl alcohol? glycerol? other? but also could be our particular GFP.
> 
> 
> Clontech has an excellent booklet on GFP technics, including some of what
> has been discussed, for reducing autofluorescence.  PBS was a suggestion.
> 
> Question:  are you trying to coverslip tissue sections or cell cultures?
> 
> Gayle Callis
> Veterinary Molecular Biology
> Montana State University
> Bozeman MT
> 
> 
> 
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 12:58:04 -0600
> From: "Munch, Barbara A" <bm27270@glaxowellcome.com>
> Subject: RE: exsanguination
> 
> Do you have a copy of the 1993 Report of the AVMA Panel on Euthanasia (JAVMA
> Vol 202, No 2, Jan 15 93 page 229-249)?  This document clearly describes
> acceptable and unacceptable methods, including sodium pentobarbital use
> (acceptable).  As long as the animal is sufficiently anesthetized to a
> surgical plane, there should be no pain felt by the animal.  Remember that
> animals may make involuntary movements not associated with pain sensation,
> staff should be fully trained to recognize the difference between a pain
> reflex and an involuntary movement.  The best reason for exsanguination is
> to remove as much blood as possible from the tisses for the best possible
> histologic sections, another reason is to collect blood samples at necropsy
> for chemistry or hematology analysis.  Many laboratories choose
> pentobarbital anesthesia/exsanguination as the most humane method of
> euthanasia for laboratory animals, and this process will work fine as long
> as staff are trained in administration and gauging the level of anesthesia.
> 
> 
> If you have any more questions, or need a copy of the Report, please let me
> know
> Barb
> Supervisor, Histo/Necropsy
> Glaxo Wellcome
> 
> 
> > -----Original Message-----
> > From:	Williams, Matthew J [PRI] [SMTP:MWillia7@prius.jnj.com]
> > Sent:	Tuesday, April 06, 1999 1:59 PM
> > To:	'HistoNet Server'
> > Subject:	exsanguination
> > 
> > 'Net folk-
> > 
> > I am looking for any documentation that explains the reasons for and
> > supports the process of exsanguination of laboratory animals before a
> > necropsy. A discussion has arisen over this subject, and the possibilty
> > that
> > it is an inhumane and unduly painful method (even though the animals are
> > anesthetized with sodium thiopental). If you care to get back to me
> > directly
> > my address is Mwillia7@prius.jnj.com.
> > 
> > Thanks!
> > 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 13:10:33 -0600
> From: Donna Carr <dkc@odsgc.net>
> Subject: PAS problems
> 
> Dear Fellow Histonetters,
>     We are currently experiencing problems with our PAS/DPAS stain,  We
> have replaced the Schiff reagent. And have tried different controls.  I
> would greatly appreciate any advice on what tissues you use as controls
> and methods of staining.  This is a recent problem that leads us to
> believe it is either a contaminated reagent or bad control.  Any input
> is welcome.   Thanks Donna
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 13:31:11 -0600
> From: "Technical Services" <techserv@dakousa.com>
> Subject: Re: Metallothionein
> 
> Hello Sarah,
> 
> We have a metallothionein antibody that is suitable for use in IHC and on
> FFPE tissues. If you would like more information please contact me.
> 
> Thank you,
> 
> Joel Weisenberger
> DAKO Corporation
> Technical Services
> 800-235-5743 x5325
> techserv@dakousa.com
> - -----Original Message-----
> From: Sarah Christo <schristo@cvm.tamu.edu>
> To: HistoNet@Pathology.swmed.edu <HistoNet@Pathology.swmed.edu>
> Date: Tuesday, April 06, 1999 11:38 AM
> Subject: Metallothionein
> 
> 
> Dear Histonetters,
>    Does anyone know of histologic techniques for a heavy metal binding
> protein called metallothionein.
> Thanks, Sarah
> 
> Sarah Christo, HT (ASCP)
> Texas A&M University
> College of Veterinary Medicine
> Dept. of Vet. Anatomy & Public Health
> College Station, TX  77868-4458
> 
> 
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 14:15:48 -0600
> From: Terri Braud <terri_braud@sfc.sch.org>
> Subject: cleaning of dispo microtome blades - reply
> 
> When we use the Tissue Tek Accu-Edge blades, we pre-clean by just
> gently wiping once with a Phlebotomy prep pak, a prepackaged 70%
> Isopropyl soaked pad. One pad will clean many blades.
> More recently, our techs seem to prefer the new Surgipath teflon
> coated blades.  They have a great edge and we do not pre-clean :-) tlb
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 14:16:12 -0600
> From: Terri Braud <terri_braud@sfc.sch.org>
> Subject: PAS
> 
> Did you check your Schiff's for reactivity?
> In a small amount of Schiff's, drop one drop of 37% formaldehyde. The
> schiffs should turn instantly, a deep rose purple. If it doesn't, then you
> know your schiffs isn't working   :-)   tlb
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 14:40:15 -0600
> From: rkline@emindustries.com
> Subject: Re: Cleaning of disposable blades
> 
> Freida,
> 
> I never cleaned them off. I used put them directly in the blade holder and
> lightly run the lead point of a pencil across the edge if they were too
> sharp.
> 
> Rande
> 
> 
> 
> 
> FreidaC@aol.com on 04/06/99 12:57:20 PM
> 
> To:   histonet@pathology.swmed.edu
> cc:
> Subject:  Cleaning of disposable blades
> 
> 
> 
> 
> I am very curious as to how many of you clean your disposable blades before
> use.  If you do clean, what do you use?
> 
> Thanks for answering my curiosity.
> 
> Freida Carson
> 
> 
> 
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 14:51:55 -0600
> From: Verrucous@aol.com
> Subject: Clean blades w/.....
> 
> Dr. Carson,
>    I clean my low profile blades w/ 100% reagent grade alcohol. Others in my 
> lab, simply score the blade against the plastic dispenser prior to use. My 
> least favorite is to store 12-20 blades at a time in the alcohol.  I feel the 
> blades stored in alcohol are less sharp.
>    I understand there is a small lubricant, between the blades at the time of 
> packaging. I like to remove it just prior to use.
> 
> Respectfully,
> 
> Bruce Gapinski HT(ASCP)
> Verrucous@aol.com 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 14:52:31 -0600
> From: Gayle Callis <uvsgc@msu.oscs.montana.edu>
> Subject: cleaning blades
> 
> Freida,
> 
> We never clean disposable blades.  Have done it in the past, but found
> it made no difference.  When I did clean them, I used acetone, instead
> of xylene or xyl substitutes.  I somehow always managed to nick the blade,
> and felt it wasn't a very safe practice.
> 
> Gayle Callis
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 15:14:56 -0600
> From: Gayle Callis <uvsgc@msu.oscs.montana.edu>
> Subject: PAS problems
> 
> How much do you reuse your Schiff's, and that test is invaluable.
> 
> Also, Culling taught me to make up the periodic acid fresh, daily!
> It is not stable forever.  We also use a coplin jar of Schiffs for a 
> week, then dipsose of it properly, and we never put working Schiffs
> back into the stock bottle (have seen that done!)  If the Schiffs is
> turning pink before use, it may also be bad, and do not freeze it.
> 
> Culling also taught us to just wash with running tap instead of sulfurous
> acid rinses, works just fine.
> 
> Gayle Callis
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 15:27:27 -0600
> From: "Sarah Christo" <schristo@cvm.tamu.edu>
> Subject: Re: Cleaning of disposable blades
> 
> Dear Freida,
>   I use alcohol and a gauze square to wipe them off before use.   -Sarah
> 
> Sarah Christo, HT (ASCP)
> Texas A&M University
> College of Veterinary Medicine
> Dept. of Vet. Anatomy & Public Health
> College Station, TX  77868-4458
> 
> >>> <FreidaC@aol.com> 04/06 11:57 AM >>>
> I am very curious as to how many of you clean your disposable blades before 
> use.  If you do clean, what do you use?
> 
> Thanks for answering my curiosity.
> 
> Freida Carson
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 15:27:55 -0600
> From: jvillalona@snet.net
> Subject: Presurecooker for ipox
> 
> I am currently using a vegetable steamer for antigent retrieval. I
> wonder how many of you are using a pressure cooker instead. what are the
> advantages of using a presure cooker, in terms of staining qualities.?
> 
> Since the cooker operates at a higher temperature; the possibilities of
> sections falling off the slides are greatly increased. Please correct me
> if iam wrong. We are looking into using a presure cooker, but we would
> like to know the pros and cons, before making a change.
> 
> Thank you in advance for your replies.
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 15:50:36 -0600
> From: marsha r price <mprice26@juno.com>
> Subject: Re: Thanks re:endogenous peroxidase, a serum blocking question
> 
> Hello Histonetters,
> I would like a recipe for making my own Primary Antibody diluent
> (Monoclonal & Polyclonal)
> Thank You.
> Marsha Price HT (ASCP), QIHC (ASCP)
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 16:02:11 -0600
> From: "Aaron Levin" <alevin@zymed.com>
> Subject: Re: Metallothionein
> 
> Dear Sarah,
> 
> Zymed Laboratories sells both a concentrated and predilute mouse monoclonal
> antibody against metallothionein. The clone is E9 and it works work with
> frozen and paraffin embedded tissue sections.
> 
> For more information please contact Zymed Laboratories technical service
> department.
> 
> Best regards,
> 
> Aaron Levin
> Zymed Laboratories
> 1-800-874-4494
> 650-871-4494 (tel)
> 650-871-4499 (fax)
> - -----Original Message-----
> From: Sarah Christo <schristo@cvm.tamu.edu>
> To: HistoNet@Pathology.swmed.edu <HistoNet@Pathology.swmed.edu>
> Date: Tuesday, April 06, 1999 11:33 AM
> Subject: Metallothionein
> 
> 
> Dear Histonetters,
>    Does anyone know of histologic techniques for a heavy metal binding
> protein called metallothionein.
> Thanks, Sarah
> 
> Sarah Christo, HT (ASCP)
> Texas A&M University
> College of Veterinary Medicine
> Dept. of Vet. Anatomy & Public Health
> College Station, TX  77868-4458
> 
> 
> 
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 17:00:30 -0600
> From: "R.Wadley" <s9803537@pop3.unsw.edu.au>
> Subject: Re: Cleaning of disposable blades
> 
> 	Dear Freida,
> 
> 	The first thing I was taught about disposable blades was that they must be
> cleaned before use.  Cleaning is not an elaborate procedure, a simple dip
> in a 100 ml specimen container of absolute ethanol, & a careful wipe with
> an ordinary facial tissue.  In the last lab I ran I used Sturkey polymer
> coated low profile blades, they come with an instruction to rinse in
> ethanol before use.
> 
> 	Why? in every case a cleaned blade will cut thinner, smoother, cleaner
> sections than one that has been used without cleaning. 
> 
> 	The only exception to this rule is the blades I used in my cryostat, there
> I used Sturkey gold coated high profile blades straight from the box.  They
> cut beautifully from ~3 - 60 um everthing from gels to bone & sections of
> entire mice.
> 
> 	Hope this helps.
> 
> 	Regards
> 
> 	Rob W.
> 
> At 12:57 PM 4/6/99 -0400, you wrote:
> >I am very curious as to how many of you clean your disposable blades before 
> >use.  If you do clean, what do you use?
> 
> 
> R. Wadley, B.App.Sc, M.L.S
> Laboratory Manager
> Cellular Analysis Facility
> School of Microbiology & Immunology
> UNSW, New South Wales, Australia, 2052
> Ph (BH) 	+61 (2) 9385 3517
> Ph (AH)	+61 (2) 9555 1239
> Fax 	+61 (2) 9385 1591
> E-mail	r.wadley@unsw.edu.au
> www	http://www.unsw.edu.au/clients/microbiology/CAF.html
> 	(Under development)
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 18:00:32 -0600
> From: "Mequita Praet" <mdpraet@bellsouth.net>
> Subject: Air Monitoring Company
> 
> I am looking for a company to come into the lab & monitor our ventilation
> system. Does anyone have any suggestion? Please send company address or
> phone number. 
> Thank you for your help.
> 
> Mequita Praet
> Dermatology Associates
> 445 S. Federal Hwy.
> Boca Raton, FL  33432
> email mdpraet@Bellsouth.net
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 18:30:38 -0600
> From: "tylee" <tylee@itis.com>
> Subject: Re: BrdU and FrdU
> 
> Dr. Miyagishima,
> The bromo and fluoro-modified nucleotides have been used to do kinetic
> studies. There are mAb which can distinguish between these two halogenated
> nucleotides (IU3 and IU4 I think, originally developed by a group in
> California system). For instance, you can inject the Br derivative into a
> patient, then after some treatment, inject the  fluoronated nucleoside
> and... after a lag period, tumor biopsies can be differentially stained to
> indicate the effectiveness of the therapy on S-phase index. Again, my memory
> of names fails me; but , I believe a woman at Rush in Chicago has published
> some of this work several years ago.
> Ty Lee
> 
> 
> - -----Original Message-----
> From: Masayuki Miyagishima, MD <mmiyagis+@pitt.edu>
> To: Histonet@Pathology.swmed.edu <Histonet@Pathology.swmed.edu>
> Date: Monday, April 05, 1999 10:45 PM
> Subject: BrdU and FrdU
> 
> 
> >In some literature, they are injecting both BrdU and FrdU.  Do we need
> >this FrdU?
> >--
> >Masayuki Miyagishima, MD
> >University of Pittsburgh,
> >Department of Surgery
> >412-647-2345, and ask the hospital operator to page 3228, please.
> >
> >
> 
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 18:31:10 -0600
> From: "Sheldon L. Brown" <slbphilip@juno.com>
> Subject: IHC On fresh loose sections
> 
> Has anyone done IHC on vibrotome sections of fresh tisuues?
> If so, is there a procedure that you would like to share?
> Please include section thickness and incubation times for the
> steps involved.
> 
> Please respond directly to my e-mail address.
> Thanks.
> 
> ___________________________________________________________________
> You don't need to buy Internet access to use free Internet e-mail.
> Get completely free e-mail from Juno at http://www.juno.com/getjuno.html
> or call Juno at (800) 654-JUNO [654-5866]
> 
> 
> ----------------------------------------------------------------------
> 
> Date: 6 Apr 1999 20:16:04 -0600
> From: Katri Tuomala <katri@istar.ca>
> Subject: Re: PAS problems
> 
> Donna Carr wrote:
> > 
> > Dear Fellow Histonetters,
> >     We are currently experiencing problems with our PAS/DPAS stain,  We
> > have replaced the Schiff reagent. And have tried different controls.  I
> > would greatly appreciate any advice on what tissues you use as controls
> > and methods of staining.  This is a recent problem that leads us to
> > believe it is either a contaminated reagent or bad control.  Any input
> > is welcome.   Thanks Donna
> 
> Hi Donna,
> We use autopsy liver tissue as a control. Just verify the presence of
> glycogen with a known control before use, not all liver tissues have
> detectable amount of glycogen present.
> 
> Katri
> 
> 
> Here are the messages received yesterday!
> 
>