date:20080911

[Histonet] Re: paraformaldehyde grade

2008-09-11 Thread David A. Wright

Hi Mary, René, & Histonet

I agree with René completely about concentration & different
paraformaldehyde grades but you might want to consider a
couple of other issues too:
a) physical state.  A coarse prill is much easier to weigh out
without contamination than a fine powder.
b) filtration.  Before perfusing the brains you have to filter
out anything that might clog the capillaries. There's usually
a milky haze left after dissolving the paraformaldehyde which
you have to remove.  I imagine that a lower grade might have
more undissolved matter, and you have to change filters a lot
even with good stuff (approx every half liter).
-David
--
David A. Wright PhD
University of Chicago Section of Neurosurgery

 Original message 
>Date: Thu, 11 Sep 2008 12:09:50 -0500 (CDT)
>From: [EMAIL PROTECTED]  
>Subject: Histonet Digest, Vol 58, Issue 13 Message: 1
>Date: Wed, 10 Sep 2008 10:10:42 -0700 (PDT)
>From: Rene J Buesa <[EMAIL PROTECTED]>
>Subject: Re: [Histonet] Paraformaldehyde
>To: histonet@lists.utsouthwestern.edu, "Mary P. Brownson"
>   <[EMAIL PROTECTED]>

>
>It does not really matter, BUT it will have to be taken into
consideration when calculating the final concentration you
want to use.
>René J.
>
>--- On Tue, 9/9/08, Mary P. Brownson <[EMAIL PROTECTED]> wrote:
>
>From: Mary P. Brownson <[EMAIL PROTECTED]>
>Subject: [Histonet] Paraformaldehyde
>To: histonet@lists.utsouthwestern.edu
>Date: Tuesday, September 9, 2008, 4:50 PM
>
>Hello,
>
>I plan to use paraformaldehyde for perfusion of rat brains. 
My question
>is; what grade paraformaldehde should I be using?  Reagent
grade, 95% or
>other?  Or does it not matter?
>

>Thank you,

>Mary Brownson
>School of Pharmacy
>University of Wyoming
>Laramie, WY 82070
>[EMAIL PROTECTED]

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[Histonet] Re: BrdU

2008-09-11 Thread David A. Wright

Hi Joost & the Histonet

I have some general comments on BrdU that i hope help.  Maybe
someone else has more specific ideas relating to your tissue 
too?  

It seems you are having problems with 2 things at once: 
a) switching species
b) switching antibodies
My guess is you only have control over the latter but either
could be the problem! 
a) species - since DNA is DNA, you don't have to worry about
species specificity of your antibody.  Unless you are doing
tissue culture, it's not however always simple to get the same
BrdU incorporation in different species.  I take it that
someone else - possibly different people - labelled the mice
and rats and also that it was done in vivo.  Even repeat
experiments on the same species are variable.  The BrdU
doesn't always go into solution equally (it needs a dilute
base, not buffered saline) and it has to have a phosphate
added on inside the cell before it is incorporated.  So
metabolic rate is important not just dose/kg.  It's a good
idea to collect control tissue from every animal to confirm
incorporation - pick any abundant, proliferating tissue.  I've
often used gut.  You then also have lots of material to work
up and compare different antibodies.  I'm probably not the
only one who'll say you have to try your 2 antibodies side by
side on the same specimen.

b) Different antibodies.  Hmmm.  You mention fixation
procedures but don't say much about what you did to recover
antigenicity in your paraffin sections before staining.  With
BrdU you have to do something just to make it accessible. In
Eukaryotes, (BrdU-containing) nuclear DNA is highly compacted
into nucleosomes with lots of histones and other proteins all
around it and then squashed down a whole lot more.  Fixation
will lock all these proteins together and make it hard for the
Ab to find its target.  I think it's a truism that all nuclear
antigens, including proteins, need some kind of harsh
treatment (heat, strong acid or base, proteinase) to make the
nucleus accessible.  It's interesting that your unconjugated
MAb worked but not the larger conjugated one - maybe it's just
penetration.
On this point, you didn't say whether your protocol was
exactly the same for each Ab.  I've used a bunch of different
anti-BrdU Abs and the manufacturers have quite different
recommendations about pretreatment - 2N HCl, formamide, SSC,
NaOH, proteinase.  If you've only got a tiny bit of tissue,
you MUST do exactly what the manufacturer says [at least the
first time] even if that's different from what worked for you
before with a different Ab (I'd do both!).  It often helps NOT
to block, so the Ab can hit the naked DNA, however exposed.

[c) time.  You mentioned different delays before processing. 
BrdU staining is one of the few things where that won't
matter. As DNA doesn't degrade like proteins, antigenicity
isn't lost with time.] 

Try Again!  Since (same logic) the DNA will still be there on
your slides, IF BrdU was actually incorporated (see a),
there's a very high likelihood that your negative slides WILL
stain with either a different antibody and/or harsher
pretreatment as recommended by the manufacturer.  I liked BD
Bioscience's MAb (clone B44) which only needed a few  minutes
in 0.07N NaOH to unmask to BrdU.  I think it's still available.

Good luck!
-David

David A. Wright PhD
University of Chicago Section of Neurosurgery

 Original message 
Histonet Digest, Vol 58, Issue 13 Message: 11
Date: Thu, 11 Sep 2008 12:04:13 +0200
From: "Bruijntjes, J.P. (Joost)" <[EMAIL PROTECTED]>
Subject: [Histonet] BrdU

Hi Histonetters

I do have a problem with BrdU, and I hope someone of you can
give a kind of solution/explanation.

I work with NALT (Nose Associated Lymphoid Tissue) of both
rats and mice, which were fixed in a fixative composed of
acetic acid, formaldehyde, ethanol and aqua dest. After 48
hours this fixative was replaced by ethanol. Later on the
tissues were dehydrated and embedded in paraffin. Just because
the lymphoid tissues material is so small, about 10 paraffin
slides were collected.

I started with the rat NALT's. One paraffin slide was stained
with HE, and a few other slides were stained with a monoclonal
antibody directed against BrdU. I was satisfied with the
staining, so far no problem.

After the rats, the same story with the mice NALT's. But I did
not get any positive staining. Some days later I repeated the
first try-out included with some positive controls from the
rat-study with the primary antibody I used in the first part,
and the biotin conjugated primary antibody as well, but again
no positive staining. The pre-treatment of the slides for rat
and mice is the same. The only difference lies in the primary
and secondary antibody.

For rat tissue I use a non-conjugated monoclonal antibody,
followed by HRP-conjugated powervision.

For mice tissue I use a biotin conjugated monoclonal antibody,
followed by a HRP-conjugated streptavidin.

Can anyone give me an explanation? The storage of

RE: [Histonet] Quantifying lipid in rat liver

2008-09-11 Thread Tony Henwood

You could try this:

Linoleic Acid Method for the demonstration of Lipids in Paraffin
Sections

Tracy & Walia (2002) have described an ingenious method to fix lipids
for staining fat embolism in paraffin sections:

(Tracy, R.E., Walia, P., (2002) "A Method to fix lipids for staining fat
embolism in paraffin sections" Histopath 41:75-79)

Solutions:

1.  Saturated solution of linoleic acid in 70% ethylene glycol
To 500ml of 70% ethylene glycol, add 5g linoleic acid and 2g lecithin,
mix for one hour and stand for several hours. Draw off the lower phase
in a separatory funnel

2.  2% chromic acid

3.  70 % ethanol

4.  5% Sodium bicarbonate

Method:
1.  Place 2mm thick sections in solution 1 for 3 days at 56oC
2.  Rinse for 8 hours in 70% ethanol
3.  Rinse in water for 8 hours
4.  Immerse in 2% chromic acid for 24 hours at 4oC.
5.  Rinse in water for 24 hours
6.  Place in 5% sodium bicarbonate for 24 hours
7.  Rinse in water 8 hours
8.  Process from 70% ethanol via ethanol and xylene to wax.
9.  Cut paraffin sections and stain with the usual fat stains (eg
oil red O) and mount in an aqueous mountant.


Regards

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
Tel: 612 9845 3306
Fax: 612 9845 3318
the children's hospital at westmead 
Cnr Hawkesbury Road and Hainsworth Street, Westmead 
Locked Bag 4001, Westmead NSW 2145 




-Original Message-
From: [EMAIL PROTECTED]
[mailto:[EMAIL PROTECTED] On Behalf Of David
Burk
Sent: Friday, 12 September 2008 12:26 AM
To: histonet@lists.utsouthwestern.edu; [EMAIL PROTECTED]
Subject: [Histonet] Quantifying lipid in rat liver 


Oh esteemed experts who have no peer,

I am in need of advice on how to quickly and easily quantify lipid
content in rat liver.  I believe a traditional approach is to
process/embed in paraffin/section/H&E and then look for the holes left
by the lipid droplets.  While this is certainly doable it does require
that your samples 'look nice'.  Unfortunately, many of the sections we
have seen here are kind of banged up and morphology is poor.

I was wondering if you all have another method that you prefer for this
type of analysis.  For example, can I just cryosection and stain with
Bodipy or Oil Red-O?  In those cases I could use the fluorescent nature
of the probe to quantify fat content very quickly on a computer.  Here,
damage caused during sectioning should not create too many problems in
quantification as the stain is specific for lipid.

If we wished to avoid image analysis altogether, could we just drop a
rat liver in an NMR and get content that way?

Thanks so much for all your help!

David Burk

Pennington Biomedical Research Center
Baton Rouge, LA 70808

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[Histonet] San Diego Chapter of CSH field trip

2008-09-11 Thread James Watson

The San Diego chapter of the Ca. Society of Histotechnology is
sponsoring a tour of all the labs at the Beckman Research Center
(Genetics and Microdiagnostic labs) adjacent to the Wild Animal Park in
Escondido:  Thursday September 25, 2005 2p.m. 

 

This tour is at no cost and should last about an hour. This tour will
not involve admission to the Wild Animal Park. So if you want to go to
Wild Animal Park you may do that on your own.

 

This trip is open to members of the chapter and other interested adults.
We can accommodate no more than thirty people.  Make you reservation
early so you will not miss out on this.

 

Parking is limited so carpool if you can.  We will need to know how many
cars are going so when you RSVP let Kris know.

 

Traveling east on 78, go past the main entrance and then make the next
left. There is a closed gate.  To gain excess, press the button on the
keypad and identify yourself. 

 

http://maps.google.com/maps?f=q&hl=en&geocode=&q=Wild+animal+park,+escon
dido,+ca&ie=UTF8&ll=33.086076,-117.039928&spn=0.064578,0.109863&z=13&iwl
oc=A

 

The cut off date and time for reservations are at 4pm Monday September
22, 2008. 

R.S.V.P.   [EMAIL PROTECTED]

 

If you are interested in becoming a member of the San Diego chapter,
please contact me.

 

James Watson HT  ASCP

Facilities Manager of Histology

GNF  Genomics Institute of the Novartis Research Foundation

Tel858-332-4647

Fax   858-812-1915  

[EMAIL PROTECTED]

 

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RE: [Histonet] bottom of water bath

2008-09-11 Thread Pamela Marcum

Margaret,

Look for a silicone microwave oven mat and cut it to size.  They come in
colours and should lay nicely on the bottom.  If need be it could be
weighted down along the sides with fishing weights or even a small metal bar
cut to size.  It is easier than painting and makes cleaning a breeze as it
lifts out and rinse clean when the water bath is emptied.  

Pamela A Marcum
University of Pennsylvania 
School of Veterinary Medicine
Comparative Orthopedic Laboratory (CORL)
382 W Street Rd
Kennett Square PA 19438
610-925-6278

-Original Message-
From: [EMAIL PROTECTED]
[mailto:[EMAIL PROTECTED] On Behalf Of Geoff
McAuliffe
Sent: Thursday, September 11, 2008 4:18 PM
To: DiCarlo, Margaret
Cc: [EMAIL PROTECTED]
Subject: Re: [Histonet] bottom of water bath

You can paint stainless steel if you prepare the surface and prime 
correctly. Call the help line of Benjamin Moore, Sherman-Williams, 
Zinsser or ?? and tell them what you need.

Geoff

DiCarlo, Margaret wrote:
> Hello Histonetters,
>
>  
>
> Working in a Bone Lab, I cut large sections which often require using 5
> X 7 inch slides.  I finally found a new water bath which is deep and
> wide enough for my lab needs.  Unfortunately, the inside of the water
> bath is stainless steel which means silver in color.  Since it can not
> be painted, does anyone have any suggestions as to what I can do or use
> to make the bottom of the water bath black so I can see the tissue
> easier?  I would appreciate your help.
>
>  
>
> Thank you. 
>
> Peggy DiCarlo HT (ASCP)
>
> Orthopedic Bone Lab
>
> Buffalo General Hospital
>
> 100 High St.
>
> Buffalo, NY  14203
>
> 716-859-1293
>
>  
>
>
>
> 2007 Best Places to Work Finalist 
> Visit our careers page at www.kaleidahealth.org/careers
>
> CONFIDENTIALITY NOTICE: This email transmission and any documents, files,
or previous e-mail messages attached to it are confidential and intended
solely for the use of the individual or entity to whom they are addressed.
If you are not the intended recipient, or a person responsible for
delivering it to the intended recipient, you are hereby notified that any
further review, disclosure, copying, dissemination, distribution, or use of
any of the information contained in or attached to this e-mail transmission
is strictly prohibited. If you have received this message in error, please
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delete all electronic files of the message. If you are unable to contact the
sender or you are not sure as to whether you are the intended recipient,
please e-mail [EMAIL PROTECTED] or call (716) 859-. 
> ___
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>
>
>   


-- 
--
**
Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583 
[EMAIL PROTECTED]
**



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Re: [Histonet] bottom of water bath

2008-09-11 Thread Geoff McAuliffe

You can paint stainless steel if you prepare the surface and prime
correctly. Call the help line of Benjamin Moore, Sherman-Williams,
Zinsser or ?? and tell them what you need.

Geoff

DiCarlo, Margaret wrote:

Hello Histonetters,

Working in a Bone Lab, I cut large sections which often require using 5
X 7 inch slides. I finally found a new water bath which is deep and
wide enough for my lab needs. Unfortunately, the inside of the water
bath is stainless steel which means silver in color. Since it can not
be painted, does anyone have any suggestions as to what I can do or use
to make the bottom of the water bath black so I can see the tissue
easier? I would appreciate your help.

Thank you.

Peggy DiCarlo HT (ASCP)

Orthopedic Bone Lab

Buffalo General Hospital

100 High St.

Buffalo, NY 14203

716-859-1293

2007 Best Places to Work Finalist
Visit our careers page at www.kaleidahealth.org/careers

CONFIDENTIALITY NOTICE: This email transmission and any documents, files, or previous e-mail messages attached to it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, or a person responsible for delivering it to the intended recipient, you are hereby notified that any further review, disclosure, copying, dissemination, distribution, or use of any of the information contained in or attached to this e-mail transmission is strictly prohibited. If you have received this message in error, please notify the sender immediately by e-mail, discard any paper copies, and delete all electronic files of the message. If you are unable to contact the sender or you are not sure as to whether you are the intended recipient, please e-mail [EMAIL PROTECTED] or call (716) 859-.
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--
**
Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583
[EMAIL PROTECTED]

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[Histonet] RE: Histonet Digest, Vol 58, Issue 13 - RECYCLERS

2008-09-11 Thread Kay, Karen


Hello,
We have used the CBG product for a few years now and are very happy with
both the equipment and support/service.  The recycler is simple to use.
We have been able to fit it into the "tasks" of the day. It has been a
very reliable instrument. The staff are always willing to help whenever
we have called them.
Karen J Kay, MLT
Pathology Supervisor
Laboratory
Chinook Regional Hospital
Lethbridge, Alberta, Canada


Message: 13
Date: Wed, 10 Sep 2008 11:58:09 -0500
From: "Bauer, Karen" <[EMAIL PROTECTED]>
Subject: [Histonet] Recyclers
To: 
Message-ID:

<[EMAIL PROTECTED]>
Content-Type: text/plain;   charset="iso-8859-1"

Hello,
 
I'm in the market to demo some recyclers.  Any vendors are welcome to
contact me personally.  I've already contacted CBG and BR Instruments,
so vendors from those areas need not reply to this message.
 
Anyone on Histonet that would like to give me their likes and dislikes
of certain recyclers, I'm all ears.
 
Thanks much,
 
Karen
 
Karen L. Bauer HT(ASCP)
Department of Pathology
Histology Section Chief
Luther Hospital
Eau Claire, WI
715-838-3205


This communication is intended for the use of the recipient to which it is 
addressed, and may contain confidential, personal and or privileged 
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recipient.  Do not copy, distribute or take action relying on it. Any 
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[Histonet] bottom of water bath

2008-09-11 Thread DiCarlo, Margaret

Hello Histonetters,

 

Working in a Bone Lab, I cut large sections which often require using 5
X 7 inch slides.  I finally found a new water bath which is deep and
wide enough for my lab needs.  Unfortunately, the inside of the water
bath is stainless steel which means silver in color.  Since it can not
be painted, does anyone have any suggestions as to what I can do or use
to make the bottom of the water bath black so I can see the tissue
easier?  I would appreciate your help.

 

Thank you. 

Peggy DiCarlo HT (ASCP)

Orthopedic Bone Lab

Buffalo General Hospital

100 High St.

Buffalo, NY  14203

716-859-1293

 



2007 Best Places to Work Finalist 
Visit our careers page at www.kaleidahealth.org/careers

CONFIDENTIALITY NOTICE: This email transmission and any documents, files, or 
previous e-mail messages attached to it are confidential and intended solely 
for the use of the individual or entity to whom they are addressed. If you are 
not the intended recipient, or a person responsible for delivering it to the 
intended recipient, you are hereby notified that any further review, 
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information contained in or attached to this e-mail transmission is strictly 
prohibited. If you have received this message in error, please notify the 
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Re: [Histonet] Pig endothelium

2008-09-11 Thread Mark Tarango

Are you putting the tissue into media like RPMI or DMEM to keep it viable
until you use it?

P.S.   Joyce, I loved the lipstick comment hehe

On Thu, Sep 11, 2008 at 11:42 AM, Alistair Lindsay <
[EMAIL PROTECTED]> wrote:

> Dear All,
>
> Does anyone have any experience with staining pig endothelium, specifically
> keeping rings of blood vessels alive?
>
> I am collecting fresh tissue from the slaughterhouse and incubating it with
> tumour necrosis factor to induce VCAM-1 expression.  However, I suspect
> that
> the tissue quickly loses viability and this is why I can¹t visualise any
> VCAM with IHC.
>
> Does anyone have any tips for keeping the rings of tissue
> going/fixing/antigen retrieval?
>
> Many thanks
>
> Alistair Lindsay MBChB MRCP
>
> Tel.: 07989404808
>
> [EMAIL PROTECTED]
> [EMAIL PROTECTED]
>
>
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Re: [Histonet] Unstained slide storage

2008-09-11 Thread Rene J Buesa

Decreased antigenicity applies to any section stored and unprotected from 
oxygen (air) oxidation.
René J.

--- On Thu, 9/11/08, [EMAIL PROTECTED] <[EMAIL PROTECTED]> wrote:

From: [EMAIL PROTECTED] <[EMAIL PROTECTED]>
Subject: [Histonet] Unstained slide storage
To: histonet@lists.utsouthwestern.edu
Date: Thursday, September 11, 2008, 2:27 PM

Daniel,
I have found that saving slides with FFPE skin sections 5-10 micrometer 
thick, the antigenicity declines with age, even when stored in the fridge. 
I have tested slides from the same block freshly cut versus 2-3 years 
stored on a microscope slide in the fridge. Whether this applies to needle 
biopsies of tumor tissue I don't know...now I just section what I need and 
return the block to the fridge.
Mel

Melville B. Vaughan, Ph. D.
Assistant Professor of Biology
Campus Coordinator, NSF Sure-Step program
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
phone 405-974-5725

Message: 15
Date: Thu, 11 Sep 2008 10:40:21 -0500
From: "Daniel Schneider" <[EMAIL PROTECTED]>
Subject: [Histonet] Protocol: Saving unstained slides...
To: histonet@lists.utsouthwestern.edu
Message-ID:
 <[EMAIL PROTECTED]>
Content-Type: text/plain; charset=ISO-8859-1

I thought I'd poll the histonet community on an issue we're facing:

On needle biopsies for tumor, what, if any, is your protocol
for making unstained slides "on the front end" for potential use with
immunostains later (so that the specimen isn't exhausted producing levels 
x
3, and you have good sections of tumor for immunostains)?

Is this protocol followed on all needle biopsies for tumor?

And what happens to any unused, unstained slides?  Are they discarded (if
so, after how long?) or kept indefinitely?

Thanks for your input!
Daniel Schneider

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[Histonet] Saving unstained slides

2008-09-11 Thread Gagnon, Eric

We pick up 10 sections on any needle/core biopsy of suspected tumour/lymphoma.  
Usually if immunostains are ordered, the request comes the same day our routine 
slides go out, or possibly the next day.  Cutting them "up front" seems the 
prudent way to go if there is any risk of the biopsy being cut away during 
subsequent sectioning for immunostains.  How about 1 stained and 3-5 unstained 
at each level?  This has saved us precious tissue many times, especially when 
multiple markers are requested.  If more markers are requested than there are 
unstained sections, the pathologist will priorize his request and we then cut 
more slides, trying to conserve as much tissue as possible.
 
As for retaining these unstained slides, they're definitely worth keeping until 
after the dx. is made, after that it's up to you how long you want to retain 
them in your protocol.  Antigenicity and storage capacity/cost are among issues 
to be considered.
 
As Greg mentioned, this is from a Canadian viewpoint.
 
Eric Gagnon MLT
Histology Laboratory
Kingston General Hospital
Kingston, Ontario, Canada
 


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RE: [Histonet] Pig endothelium

2008-09-11 Thread Weems, Joyce

Lipstick, maybe? Sorry, couldn't help it... j

-Original Message-
From: [EMAIL PROTECTED] [mailto:[EMAIL PROTECTED] On Behalf Of Alistair Lindsay
Sent: Thursday, September 11, 2008 2:43 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Pig endothelium

Dear All,

Does anyone have any experience with staining pig endothelium, specifically 
keeping rings of blood vessels alive?

I am collecting fresh tissue from the slaughterhouse and incubating it with 
tumour necrosis factor to induce VCAM-1 expression.  However, I suspect that 
the tissue quickly loses viability and this is why I can¹t visualise any VCAM 
with IHC. 

Does anyone have any tips for keeping the rings of tissue going/fixing/antigen 
retrieval?

Many thanks

Alistair Lindsay MBChB MRCP

Tel.: 07989404808

[EMAIL PROTECTED]
[EMAIL PROTECTED]

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[Histonet] Pig endothelium

2008-09-11 Thread Alistair Lindsay

Dear All,

Does anyone have any experience with staining pig endothelium, specifically
keeping rings of blood vessels alive?

I am collecting fresh tissue from the slaughterhouse and incubating it with
tumour necrosis factor to induce VCAM-1 expression.  However, I suspect that
the tissue quickly loses viability and this is why I can¹t visualise any
VCAM with IHC. 

Does anyone have any tips for keeping the rings of tissue
going/fixing/antigen retrieval?

Many thanks

Alistair Lindsay MBChB MRCP

Tel.: 07989404808

[EMAIL PROTECTED]
[EMAIL PROTECTED]


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[Histonet] Unstained slide storage

2008-09-11 Thread MVaughan4

Daniel,
I have found that saving slides with FFPE skin sections 5-10 micrometer 
thick, the antigenicity declines with age, even when stored in the fridge. 
I have tested slides from the same block freshly cut versus 2-3 years 
stored on a microscope slide in the fridge. Whether this applies to needle 
biopsies of tumor tissue I don't know...now I just section what I need and 
return the block to the fridge.
Mel


Melville B. Vaughan, Ph. D.
Assistant Professor of Biology
Campus Coordinator, NSF Sure-Step program
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
phone 405-974-5725

Message: 15
Date: Thu, 11 Sep 2008 10:40:21 -0500
From: "Daniel Schneider" <[EMAIL PROTECTED]>
Subject: [Histonet] Protocol: Saving unstained slides...
To: histonet@lists.utsouthwestern.edu
Message-ID:
 <[EMAIL PROTECTED]>
Content-Type: text/plain; charset=ISO-8859-1

I thought I'd poll the histonet community on an issue we're facing:

On needle biopsies for tumor, what, if any, is your protocol
for making unstained slides "on the front end" for potential use with
immunostains later (so that the specimen isn't exhausted producing levels 
x
3, and you have good sections of tumor for immunostains)?

Is this protocol followed on all needle biopsies for tumor?

And what happens to any unused, unstained slides?  Are they discarded (if
so, after how long?) or kept indefinitely?

Thanks for your input!
Daniel Schneider


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Re: [Histonet] Protocol: Saving unstained slides...

2008-09-11 Thread Greg Dobbin



Hi Daniel,
All of our "blanks" are saved for a short time (ie 2-4 weeks) just in
case, but those from cases that have very little tissue left in the
block or small malignant foci are filed with the H&E slides (and any
other stained slides of course) that go with the case. The idea is that
if in 2 years we want to restain the tumour for something new or (heaven
forbid!) a re-check, the pathologist would generally be asking to see
the original H&E's anyway; and at that time we would discover that
indeed blanks do exist on the case already.
Hope this helps (Disclaimer-I'm in Canada and none of this response
necessarily addresses either CAP or CLIA regulations).
Cheers!
Greg

Greg Dobbin, R.T.
Chief Technologist, Anatomic Pathology
Dept. of Laboratory Medicine,
Queen Elizabeth Hospital,
P.O. Box 6600
Charlottetown, PEC1A 8T5
Phone: (902) 894-2337
Fax: (902) 894-2385

"Success is not the key to happiness. 
Happiness is the key to success. 
If you love what you are doing, 
you will be successful." 
- Albert Schweitzer


Greg Dobbin, R.T.
Chief Technologist, Anatomic Pathology
Dept. of Laboratory Medicine,
Queen Elizabeth Hospital,
P.O. Box 6600
Charlottetown, PEC1A 8T5
Phone: (902) 894-2337
Fax: (902) 894-2385

"Success is not the key to happiness. 
Happiness is the key to success. 
If you love what you are doing, 
you will be successful." 
- Albert Schweitzer


>>> "Daniel Schneider" <[EMAIL PROTECTED]> 9/11/2008 12:40 PM >>>
I thought I'd poll the histonet community on an issue we're facing:

On needle biopsies for tumor, what, if any, is your protocol
for making unstained slides "on the front end" for potential use with
immunostains later (so that the specimen isn't exhausted producing
levels x
3, and you have good sections of tumor for immunostains)?

Is this protocol followed on all needle biopsies for tumor?

And what happens to any unused, unstained slides?  Are they discarded
(if
so, after how long?) or kept indefinitely?

Thanks for your input!
Daniel Schneider
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[Histonet] formalin-formula and osmolarity

2008-09-11 Thread Gudrun Lang

Hi all,

I need the help of those with a talent in chemical calculation.

This formula is used in our institute since 1970, but nobody knows the
origin or a publication.

 

Formalin-formula: 8% buffered Formaldehyd

2 Liter 36% Formaldehyd

2 Liter Phosphatpuffer pH 7,8

6 Liter Aqua dest.

EndpH 7,4

 

commercial Phosphatpuffer contents:

Natrium di-hydrogenphosphat monohydrat  1,810 g 

di-Natrium monohydrogenphosphat dihydrat3,756 g 

Natriumchlorid  4,4 g 

Water ad  1000
ml

 

Phosphatebuffer has a molarity of  34,2 mmol/l.

 

Can anybody calculate the osmolarity of the ready formalin-solution for me? 

What could be the advantages or disadvantages compared to Lillies 4% NBF or
Carson's 4% NBF?

Is the buffer-effect big enough in comparison to other formulas with 0,1 M
Phosphatebuffer?

 

Pleas help, any input is appreciated.

Gudrun Lang

 

Linz, Austria

.

 

 

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[Histonet] test: please delete

2008-09-11 Thread Bartlett, Jeanine (CDC/CCID/NCZVED)




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[Histonet] Protocol: Saving unstained slides...

2008-09-11 Thread Daniel Schneider

I thought I'd poll the histonet community on an issue we're facing:

On needle biopsies for tumor, what, if any, is your protocol
for making unstained slides "on the front end" for potential use with
immunostains later (so that the specimen isn't exhausted producing levels x
3, and you have good sections of tumor for immunostains)?

Is this protocol followed on all needle biopsies for tumor?

And what happens to any unused, unstained slides?  Are they discarded (if
so, after how long?) or kept indefinitely?

Thanks for your input!
Daniel Schneider
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[Histonet] Quantifying lipid in rat liver

2008-09-11 Thread David Burk

Oh esteemed experts who have no peer,

I am in need of advice on how to quickly and easily quantify lipid
content in rat liver.  I believe a traditional approach is to
process/embed in paraffin/section/H&E and then look for the holes left
by the lipid droplets.  While this is certainly doable it does require
that your samples 'look nice'.  Unfortunately, many of the sections we
have seen here are kind of banged up and morphology is poor.

I was wondering if you all have another method that you prefer for this
type of analysis.  For example, can I just cryosection and stain with
Bodipy or Oil Red-O?  In those cases I could use the fluorescent nature
of the probe to quantify fat content very quickly on a computer.  Here,
damage caused during sectioning should not create too many problems in
quantification as the stain is specific for lipid.

If we wished to avoid image analysis altogether, could we just drop a
rat liver in an NMR and get content that way?

Thanks so much for all your help!

David Burk

Pennington Biomedical Research Center
Baton Rouge, LA 70808

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[Histonet] Antigen retrieval 'tween night and next day