[Histonet] Tunel

[Amos Brooks via Histonet]

Hi,

> I hand stain this. It is a finicky test and benefits from a personal
touch. When I have a ton of them, sometimes I do the enzyme on the stainer
and then take it off for the primary and secondary then DAB on the stainer.

Amos


Message: 1
Date: Fri, 19 Oct 2018 19:24:01 +
From: "Cathy M. Mathis/Comparative Medicine" 
To: "histonet@lists.utsouthwestern.edu"
    
Subject: [Histonet] TUNEL on Leica Bond
Message-ID:
<
4816f916b11aec4e92a0d01372f573e2012cfd3...@exchdb8.medctr.ad.wfubmc.edu>

Content-Type: text/plain; charset="us-ascii"

Hello Histo-friends,
Is anyone doing a TUNEL stain on any of the Leica Bond instruments?  I will
be attempting to set this up and thought I would reach out first.
Thank you,
Cathy

Cathy M. Mathis
Laboratory Technician IV
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[Histonet] TUNEL on Leica Bond

[Cathy M. Mathis/Comparative Medicine via Histonet]

Hello Histo-friends,
Is anyone doing a TUNEL stain on any of the Leica Bond instruments?  I will be 
attempting to set this up and thought I would reach out first.
Thank you,
Cathy

Cathy M. Mathis 
Laboratory Technician IV

-Original Message-
From: histonet-requ...@lists.utsouthwestern.edu 
[mailto:histonet-requ...@lists.utsouthwestern.edu] 
Sent: Friday, October 19, 2018 1:00 PM
To: histonet@lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 179, Issue 16

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Today's Topics:

   1. toluene substitute (Michelle Jamison)
   2. RUO antibody (Erin McCarthy)
   3. Re: Restaining old Heamatology smears (Eddie Martin)
   4. Re: BMP-2 protocol for bone (Eddie Martin)


--

Message: 1
Date: Thu, 18 Oct 2018 14:48:50 -0600
From: Michelle Jamison 
To: "histonet@lists.utsouthwestern.edu"

Subject: [Histonet] toluene substitute
Message-ID: 
Content-Type: text/plain; charset=utf-8; format=flowed

Hi All, I am from industry. We are interested in developing an 
instrument that stains then prepares microscope slides to be 
coverslipped. If you do coverslip, what substitute do you use instead of 
toluene to go to mounting media? Thank you! Michelle
-- 

All the best to you,

Michelle Jamison

/Bio Research Associate /

/ELITechGroup Inc. / ???370 W. 1700 S.?? ???Logan Utah ??? USA

m.jami...@elitechgroup.com  ? 
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Message: 2
Date: Thu, 18 Oct 2018 16:30:14 -0500
From: Erin McCarthy 
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] RUO antibody
Message-ID:

Content-Type: text/plain; charset="UTF-8"

Hi All,

I am looking to see if anyone has had any experience with the antibody AXL.
We are trying to work it up on our Roche Discovery Ultra and it does not
seem like it stains both nuclear and cytoplasic cell features. We only seem
to see cytoplasmic staining, and in general most tumors are showing
negative results.

I currently am using the Ultraview kit for optimization, our pathologist
likes CC2 for 44 min, Ab. Inc for 40 min using a 1:100 dilution.

Anyone have any suggestions that might help? If you want more detailed info
I can provide as well!

-- 

Erin McCarthy, HT (ASCP)
Pathology Lab Supervisor

Tempus Labs
600 W. Chicago Ave.
Chicago IL 60654
Ph:(312) 638-6344 Ext.3835

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Message: 3
Date: Thu, 18 Oct 2018 18:20:53 -0400
From: Eddie Martin 
To: "Reilly, Laurie" 
Cc: "Histonet@lists.utsouthwestern.edu"

Subject: Re: [Histonet] Restaining old Heamatology smears
Message-ID: <0e7f124e-9821-4a32-a438-63f45ad45...@gmail.com>
Content-Type: text/plain;   charset=utf-8

Hi Laurie,

Blood is considered a tissue so it is very much still Histo related. 

Regarding your smears, this also 

Re: [Histonet] TUNEL

[Teri Johnson via Histonet]

Hi Bernice,

Likely there is something in your TUNEL procedure that is causing a problem 
with histochemical nuclear staining. If it isn't the pre-treatment, it might be 
the DAPI. After all, they do bind the same structures, and the DAPI might be 
winning that competition.

Teri Johnson
Manager, Clinical Trial Testing

T +1 760 516 5954
M +1 442 222 4503
tejohn...@genoptix.com

Navigate BioPharma, Inc.
A Novartis Company
1890 Rutherford Rd.
Carlsbad, CA  92008
USA




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Re: [Histonet] TUNEL

[Caroline Miller via Histonet]

I have done that a lot and I don't see why it would not pick up hx or DAPI.
Was there anything left on the slide? Maybe it was mounted in aqueous
already??

mills

On Fri, Mar 3, 2017 at 11:42 AM, Bernice Frederick via Histonet <
histonet@lists.utsouthwestern.edu> wrote:

> Hello all.
> Would there be any reason that slides stained with TUNEL (ISH) would not
> pick up the Mayers counterstain? We use it for IHC and it is fine. Mayers
> for 7 minutes and then ammonia water for 1 minute. We do that rather than
> rinse for 15 minutes and there are usually no issues. Could a reagent in
> the stain be causing it? DAPI is used...
> Bernice
>
> Bernice Frederick HTL (ASCP)
> Senior Research Tech
> Pathology Core Facility
> Robert. H. Lurie Cancer Center
> Northwestern University
> 710 N Fairbanks Court
> Olson 8-421
> Chicago,IL 60611
> 312-503-3723
> b-freder...@northwestern.edu
>
> ___
> Histonet mailing list
> Histonet@lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>



-- 
Caroline Miller (mills)
Director of Histology
3Scan.com
415 2187297
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[Histonet] TUNEL

[Bernice Frederick via Histonet]

Hello all.
Would there be any reason that slides stained with TUNEL (ISH) would not pick 
up the Mayers counterstain? We use it for IHC and it is fine. Mayers for 7 
minutes and then ammonia water for 1 minute. We do that rather than rinse for 
15 minutes and there are usually no issues. Could a reagent in the stain be 
causing it? DAPI is used...
Bernice

Bernice Frederick HTL (ASCP)
Senior Research Tech
Pathology Core Facility
Robert. H. Lurie Cancer Center
Northwestern University
710 N Fairbanks Court
Olson 8-421
Chicago,IL 60611
312-503-3723
b-freder...@northwestern.edu

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[Histonet] Tunel with Apoptag

[Amos Brooks via Histonet]

Hi,
 I am contentedly using the Millipore (Chemicon) Apoptag IHC kit for
Tunel stains. It works great... *BUT* I have been getting requests for a
fluorescent tag rather than a chromogenic. The contents of the fluorescent
kits are identical except for the anti-Dig. In the chromogenic kit is HRP
tagged and the fluorescent is IF tagged. All I need is the IF tagged part,
but wouldn't you know it, they don't sell just that part. No, you need to
buy the whole dang kit for the one component you need. So, clearly I need
to find an alternative fluorescent tagged anti-DIG antibody that would
detect this TDT enzyme. Any suggestions?

Thanks,
Amos
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[Histonet] Tunel stain by IHC

[Coppin, Margaret]

Hello everyone,

I have a pathologist that is looking for a lab that performs the Tunel stain. I 
guess she wants to stain some research mouse samples to look for apoptosis.

Thank you,

Margaret G. Coppin, HT(ASCP)
Technical Supervisor,
Immunohistochemistry

ARUP Laboratories
500 Chipeta Way
Salt Lake City, UT 84108
phone: (801)583-2787 X3869
email: copp...@aruplab.com




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[Histonet] TUNEL on Ventana

[mesruh turkekul]

Hi Guys,

Has anyone of you done TUNEL on Ventana?

Mesruh Turkekul
mskcc.org
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[Histonet] Tunel

[Amos Brooks]

Hi,
 I love the Millipore kit S7101. The data sheet (instructions)
require a bit of distilling into a palatable form, but it works great
once you clean up the procedure some. If you need a hand with this one
drop me a line. I do it very often. Incidentally as with most kits
some parts end up used up faster than others. Millipore sells the
individual components as well so no sweat there.

Good luck,
Amos


Message: 5
Date: Fri, 3 Sep 2010 16:26:31 -0400
From: Vanessa J. Phelan vjp2...@columbia.edu
Subject: [Histonet] TUNEL
To: histonet@lists.utsouthwestern.edu
   Histonet@lists.utsouthwestern.edu
Message-ID: c8a6d237.3de2%phe...@icg.cpmc.columbia.edu
Content-Type: text/plain; charset=iso-8859-1

Hi everyone,

I am hoping you can help me I am looking for some advice on how to do
TUNEL staining (fluorescent or enzymatic), kits recommended/ protocol.
Any help would be greatly appreciated.

Thanking you in advance

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[Histonet] TUNEL

[Vanessa J. Phelan]

Hi everyone,

I am hoping you can help me I am looking for some advice on how to do TUNEL 
staining (fluorescent or enzymatic), kits recommended/ protocol. Any help would 
be greatly appreciated.

Thanking you in advance

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RE: [Histonet] TUNEL

[Liz Chlipala]

We use the Roche Kit with good success.

Liz

Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Manager
Premier Laboratory, LLC
PO Box 18592
Boulder, Colorado 80308
office (303) 682-3949 
fax (303) 682-9060
www.premierlab.com
 
 
Ship to Address:
1567 Skyway Drive, Unit E
Longmont, Colorado 80504


-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Vanessa
J. Phelan
Sent: Friday, September 03, 2010 2:27 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] TUNEL 


Hi everyone,

I am hoping you can help me I am looking for some advice on how to do
TUNEL staining (fluorescent or enzymatic), kits recommended/ protocol.
Any help would be greatly appreciated.

Thanking you in advance

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[Histonet] TUNEL

[Michelle MacVeigh-Aloni]

We also use Roche kits. The cat # 11 684 795 910 is for fluorescent and cat# 11 
684 817 910 is for loight microscope. There are aditional chemicals to 
purchase, beside the kit. 
Call Roche for more info.

Good luck!

Michelle
USC School of Medicine
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[Histonet] TUNEL of frozen thick rat brain slices.

[Guillermo Palchik]

Dear Histologists,
Has anyone done TUNEL on 40 um flash-frozen rat brain sections? I have tried 
tweaking the protocol (we use the Millipore Apoptag kit  - S7101) from our 
ongoing 20 um slices by increasing the incubation times from 1 hour to 2 and 4 
hours but we still get poor staining. Also, could I do overnight incubations? 
the tissue is flash frozen, but it does get fixed at the beginning of the 
protocol with PFA (4%) and subsequently with acetic acid-EtOH.
Thanks,
Gil
 
-- 
Guillermo Palchik
Ph.D. Candidate - Interdisciplinary Program in Neuroscience
Georgetown University Medical Center
Research Building Room W 217
3970 Reservoir Rd. NW, Washington, DC 20007
Lab: 202-687-7825


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RE: [Histonet] TUNEL of frozen thick rat brain slices.

[Montina Van Meter]

Gil,
 
I routinely incubate free-floating 40um rat brain sections overnight or
longer @ 4 degrees centigrade according to the particular antibody that
I am working with.  Our sections are perfused in 4% paraformaldehyde and
cryoprotected with 30% sucrose prior to sectioning.  How long are you
fixing the tissue prior to staining?

 Tina



-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of
Guillermo Palchik
Sent: Wednesday, August 04, 2010 11:21 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] TUNEL of frozen thick rat brain slices.

Dear Histologists,
Has anyone done TUNEL on 40 um flash-frozen rat brain sections? I have
tried tweaking the protocol (we use the Millipore Apoptag kit  - S7101)
from our ongoing 20 um slices by increasing the incubation times from 1
hour to 2 and 4 hours but we still get poor staining. Also, could I do
overnight incubations? the tissue is flash frozen, but it does get fixed
at the beginning of the protocol with PFA (4%) and subsequently with
acetic acid-EtOH.
Thanks,
Gil
 
-- 
Guillermo Palchik
Ph.D. Candidate - Interdisciplinary Program in Neuroscience
Georgetown University Medical Center
Research Building Room W 217
3970 Reservoir Rd. NW, Washington, DC 20007
Lab: 202-687-7825


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[Histonet] TUNEL

[Melissa Mazan]

Hi all,
I am trying to do an apoptosis assay on mouse lung tissue (FFPE), using 
the Trevigen (formerly Roche) TACs TdT system (fluorescent).  I am 
generating a positive control using nuclease provided by the kit - and 
get a beautiful, clean, strong signal.  Problem is, when I do the same 
assay using the actual tissues, I get no staining whatsoever.  I have 
included tissues that should have apoptotic cells, including 
hyperoxia-damaged lung tissue, neonatal tissue, and post-pneumonectomy 
tissue.  When I contact the company, they tell me that if my positive 
controls are working, then the assay is working, and there is no way to 
optomize it further.  I have already tried cytonin v. Prot K, and Mn2+ 
v. Co2+, with no improvement.  Biologically, the hyperoxia tissues ought 
to be full of apoptotic cells, and all the others should have at least 
some.  Does anyone have a good protocol or a better kit to suggest? 
Thanks! Melissa


histonet-requ...@lists.utsouthwestern.edu wrote:

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When replying, please edit your Subject line so it is more specific
than Re: Contents of Histonet digest...


Today's Topics:

   1. Ga.Society For Histotechnology Reservation Link
  (Shirley A. Powell)
   2. Re: Histonet Digest, Vol 74, Issue 34 (Rhonda Henshall-Powell)
   3. need help with stain (Perry, Margaret)
   4. Re: need help with stain (Rene J Buesa)
   5. Thomas Crowell is out of the office. (thomas.crow...@novartis.com)
   6. RE: [SPAM-HC] - [Histonet] Microtomist's Vade-Mecum on the
  Web - Email found in subject (McCormick, James)
   7. RE: Re: Diff-Quik (Tony Henwood)
   8. A blast from the past (Jeffrey Silverman)
   9. (no subject) (Green JumpyOne)
  10. honing compound (Jennifer MacDonald)
  11. RE: A blast from the past (Mike Pence)
  12. Re : KP Markers (Vanessa Gutierrez)
  13. b-5 (Vickroy, Jim)
  14. Friday Hour of Fun (Breeden, Sara)
  15. RE: b-5 (Weems, Joyce)
  16. job opening  (Konni Black)
  17. Decontamination of a Cryostat (Sharon.Davis-Devine)
  18. Re: Decontamination of a Cryostat
  (jan.mins...@leica-microsystems.com)
  19. RE: Friday Hour of Fun (Mahoney,Janice A)
  20. Kaposi's sarcoma block (Houston, Ronald)
  21. RE: Decontamination of a Cryostat (Morken, Tim)


--

Message: 1
Date: Thu, 28 Jan 2010 13:35:48 -0500
From: Shirley A. Powell powell...@mercer.edu
Subject: [Histonet] Ga.Society For Histotechnology Reservation Link
To: histonet@lists.utsouthwestern.edu
histonet@lists.utsouthwestern.edu
Message-ID:
9bf995bc0e47744e9673a41486e24ee22429f09...@mercermail.merceru.local
Content-Type: text/plain; charset=us-ascii

Hi All,

I wanted to pass this link from the hotel on to you guys who will be attending the 
Georgia Society for Histotechnology meeting March 26-28 this year.  This goes 
straight to the hotel and already has our code entered for you to reserve your rooms 
at the Evergreen Marriott Conference Resort.  For the complete program and 
registration form go to www.histosearch.com/gshhttp://www.histosearch.com/gsh 
and go to the symposium page.  You will find the hotel link there too.

See you there.

Shirley Powell
GSH Secretary



Subject: Ga.Society For Histotechnology Reservation Link

Simply cut and paste the link below and include with your electronic 
correspondence to facilitate the reservation process. Your guests will be 
directed to the property's home page with the code already entered in the 
appropriate field. All they need to do is enter their arrival date to begin the 
reservation process.





Evergreen Marriott Conference Resort 
http://www.marriott.com/hotels/travel/atleg?groupCode=gshgshaapp=resvlinkfromDate=3/26/10toDate=3/27/10





[http://www.marriott.com/Images/Brands/MCC/Logos/MCC_logowhitefield_120x60.gif]http://www.marriott.com/hotels/travel/atleg?groupCode=gshgshaapp=resvlinkfromDate=3/26/10toDate=3/27/10





http://www.marriott.com/hotels/travel/atleg?groupCode=gshgshaapp=resvlinkfromDate=3/26/10toDate=3/27/10














--

Message: 2
Date: Thu, 28 Jan 2010 19:08:33 + (GMT)
From: Rhonda Henshall-Powell rlhenshall_pow...@yahoo.co.uk
Subject: [Histonet] Re: Histonet Digest, Vol 74, Issue 34
To: histonet@lists.utsouthwestern.edu
Message-ID: 828544.67370...@web23205.mail.ird.yahoo.com
Content-Type: text/plain; charset=iso-8859-1

Dear Nick,

When I was growing and harvesting 3D cell cultures (spheroids grown in 
matrigel) we would remove the culture medium, draw up the gel in a needle-less 
syringe and 

[Histonet] TUNEL

[Sarka Lhotak]

Hi Melissa,

I use the Trevigen system with great success on mouse tissue using
biotinylated dNTPs, Streptavidin HRP and NovaREd. It should work just
as well with fluorescence. Mouse thymus is a great positive control.
My protocol is as follows:

-Xylenes, ethanols, water.

-Proteinase K 30 minutes at 37C

-Endog. peroxidase quenching: 45 ml methanol+5ml 30% H2O2, 5 min.

-1x labeling buffer, 5 min

-Reaction mixture: 1x labeling b. 50ul + TdTdNTP mix 1ul+ 50xCo2+ 1ul +
TdT enzyme 1ul. 1hr at 37C.
 
-1x Stop buffer, 5 min RT
Then I use the Streptavidin HRP and Nova Red that I use for IHC,
exactly the same way.

I found that the Endog. peroxidase quenching step was ESSENTIAL for
this reaction to work, i.e. probably doing more than just quenching,
without that there was NO reaction at all.  

Hope this helps somewhat, don't hesitate to contact me if you have more
questions. The beauty of the Trevigen system is that you can buy
individual reagents, no need to be paying big bucks for a full kit.
(No commercial interest in Trevigen :), just a happy customer).

Sarka Lhotak

McMaster University, Hamilton
 

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RE: [Histonet] TUNEL on fixed brain tissue

[Patsy Ruegg]

Human brain?  I prefer Cleaved Caspase 3 to Tunnel, in my experience Tunnel
does not distinguish apoptosis from necrosis but CC3 will only label cells
under going apoptosis.
Patsy 

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Guillermo
Palchik
Sent: Wednesday, December 24, 2008 8:06 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] TUNEL on fixed brain tissue

Dear Histologists,
Does anyone have a good protocol for doing TUNEL on fixed brain slides?
Thanks

Guillermo Palchik
g...@georgetown.edu







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[Histonet] TUNEL on fixed brain tissue

[Guillermo Palchik]

Dear Histologists,
Does anyone have a good protocol for doing TUNEL on fixed brain slides?
Thanks

Guillermo Palchik
g...@georgetown.edu







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[Histonet] TUNEL on FIXED rat PUPS

[Guillermo Palchik]

Dear Histologists...

I am currently doing an experiment in which I perform TUNEL on brain  
slices that were treated with certain drugs, to examine the levels of  
apoptotic cell death. The rat’s brain is flash frozen (scooped from  
the skull, directly into cold isopentane) air dried, and then stored  
into a -80 C freezer.  We then cut the brain into 20 um slices using a  
cryostat and mount them on slides, and store them in a -20 C freezer,  
where they remain until the TUNEL step. The TUNEL is performed using  
the Apoptag Plus kit from Chemicon using Peroxidase/DAB (Cat. # S7101)  
and counterstaining with 0.5% methyl green. This has been done in the  
lab for quite some time now and we are able to get results…
We want to switch over to perfused brains (instead of flash frozen).  
However, we have had ZERO positive staining once the brains are fixed  
(in 4% PF). This has been corroborated by other lab members that have  
tried it for a couple of years already…
The protocol that came with the kit has a section for flash frozen and  
a section for fixed tissue, and since I have done TUNEL using fixed  
tissue before, I know that it is possible to do TUNEL in fixed tissue,  
however we cannot get any positive staining whatsoever… Along these  
lines, since the first step of the Apoptag TUNEL protocol is to fix  
the tissue with 4% PF (for 10 min), this has led me to believe that  
the problem is indeed the initial fixation with PF (at the time of  
perfusion).
I should say that we work with rat PUPS for this and that the immature  
brain is not the same as the mature brain (the immature brain has more  
fat, for example) and that this might be the cause of the problem…In  
any case, I was doing some research and I wanted to try using Zinc  
Formalin as a perfusate and see if this would allow us to do the TUNEL.


I would appreciate any comments and suggestions regarding this. I am  
sorry for the lengthy email, but I wanted to show a more or less  
complete picture, in case I am overlooking other factors…


Guillermo Palchik
g...@georgetown.edu






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RE: [Histonet] TUNEL on FIXED rat PUPS

[Liz Chlipala]

 sample
26. Incubate at 37°C in the Dako Hybridizer   30 minutes

NOTE:  If necessary to ensure a homogenous spread of TUNEL
reaction mixture across the sample and to avoid evaporative loss, the
samples can be covered with a coverslip during incubation.

27. PBS pH7.4 0.1% tween rinse  3 minutes or 
more
28. PBS pH7.4 0.1% tween rinse  3 minutes or 
more
29. Wash Buffer 1 minute
30. Prepare Vulcan Fast Red Substrate Solution

To 2.5 mls of Vulcan Fast Red Buffer add one drop of vial A and one
drop of vial b.  Use immediately, discard any leftover reagent.

31. Vulcan Fast Red Substrate Solution  10 - 30 
minutes

32. Distilled water 2 minutes
33. Distilled water 2 minutes
34. Hematoxylin 3 minutes
35. Distilled water 1 minute
36. Wash Buffer 1 minute
37. Distilled water 1 minute
38. Let slides air dry
39. Xylene  1 minute
40. Mount and coverslip
41. Review positive and negative control slides for appropriate staining


Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Manager
Premier Laboratory, LLC
P.O. Box 18592
Boulder, CO 80308
phone (303) 682-3949
fax (303) 682-9060
l...@premierlab.com
www.premierlab.com

Ship to Address:

Premier Laboratory, LLC
1567 Skyway Drive
Unit E
Longmont, CO 80504

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Guillermo 
Palchik
Sent: Tuesday, December 16, 2008 3:37 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] TUNEL on FIXED rat PUPS

Dear Histologists...

I am currently doing an experiment in which I perform TUNEL on brain slices 
that were treated with certain drugs, to examine the levels of apoptotic cell 
death. The rat's brain is flash frozen (scooped from the skull, directly into 
cold isopentane) air dried, and then stored into a -80 C freezer.  We then cut 
the brain into 20 um slices using a cryostat and mount them on slides, and 
store them in a -20 C freezer, where they remain until the TUNEL step. The 
TUNEL is performed using the Apoptag Plus kit from Chemicon using 
Peroxidase/DAB (Cat. # S7101) and counterstaining with 0.5% methyl green. This 
has been done in the lab for quite some time now and we are able to get 
results... We want to switch over to perfused brains (instead of flash frozen). 
However, we have had ZERO positive staining once the brains are fixed (in 4% 
PF). This has been corroborated by other lab members that have tried it for a 
couple of years already... The protocol that came with the kit has a section 
for flash frozen and a section for fixed tissue, and since I have done TUNEL 
using fixed tissue before, I know that it is possible to do TUNEL in fixed 
tissue, however we cannot get any positive staining whatsoever... Along these 
lines, since the first step of the Apoptag TUNEL protocol is to fix the tissue 
with 4% PF (for 10 min), this has led me to believe that the problem is indeed 
the initial fixation with PF (at the time of perfusion).
I should say that we work with rat PUPS for this and that the immature brain is 
not the same as the mature brain (the immature brain has more fat, for example) 
and that this might be the cause of the problem...In any case, I was doing some 
research and I wanted to try using Zinc Formalin as a perfusate and see if this 
would allow us to do the TUNEL.

I would appreciate any comments and suggestions regarding this. I am sorry for 
the lengthy email, but I wanted to show a more or less complete picture, in 
case I am overlooking other factors...

Guillermo Palchik
g...@georgetown.edu






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