Re: [Histonet] Cold ischemic times

[E. Wayne Johnson via Histonet]

When I first saw the title of your email I thought it was
a publication OR a commentary on The Era.

New York Times
Cold Ischemic Times
Global Times
Los Angeles Times.

*

It was the worst of times
it was the best of times
Man, these are such cold ischemic times,

(Brother, can you paradigm?)

We never knew what time it was
but we knew how sublime it was.

E. Wayne Johnson DVM
Enable Ag Tech
Beijing.


Greg Dobbin via Histonet wrote:

This is more of a survey than a question:

For those of you tracking and documenting your cold ischemic times for
breast tissue (ie time out of body to time sliced [as needed] and immersed
in formalin), and I assume most of you are...*what is your average time?*

*Background:*
*I ask because my director was looking for suggestions for quality
indicators to report and while I feel like our cold ischemic average time
is impressive at ~17 mins, she says that is pretty standard for everyone.*

Thanks,
Greg




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Re: [Histonet] The LEGAL side of specimen ownership

[E. Wayne Johnson via Histonet]

 Townsend synth intro [dito 
dito dito dito nano dito nano dito]

Washington University vs WJ Catalona (2005, 2008) dealt with the subject of 
research samples
which had been formally donated to the university, not with body parts excised 
during the course of
surgery that the patient wants to take home and cook or feed to their pet or
have bronzed and display on the mantle.


Actually the use of the donation form in the case of WU vs Catalona implies that
the patient had rights over the material which were then waived by the 
completion of
the form.  The issue in "Catalona" was whether or not the researcher Catalona 
could
retain some research tissue if the donor so directeed it to stay with him, or if
the university could hang onto the stuff because it was donated to the 
university.

Portuguese national law follows a Lockean interpretation of personal property 
rights
that the body parts belong to the individual and on his death the ownership 
passes to
the family.

Nevertheless physical possession is always a large practical part of a legal 
right
and it could be some difficulty wresting the material out of the clutching hands
of biomedical bureaucrats.

https://youtu.be/UDfAdHBtK_Q?t=445


E. Wayne Johnson DVM
Enable AgTech
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Terri Braud via Histonet wrote:

Sorry, E. Wayne, but in the USA, according to December 2004 JAMA The Journal of 
the American Medical Association 292(20):2500-5, recent examination of these 
issues by a US federal court resulted in a ruling that individuals do not 
retain rights of ownership or control of biological materials.  It belongs to 
the receiving laboratory.
A small collection of case law has determined that samples are controlled and 
owned not by those who contributed them but by researchers or their 
institutions. Taken together, the cases do not offer clear guidance; they are 
consistent only in their denial of a right claimed by individuals who 
contributed samples. Genet Med. 2011 Jun; 13(6): 569-575.
It is not YOUR gallbladder if you go to a hospital to have it removed. It 
becomes the property of the hospital or where ever they chose to send it.
 DONG DONG
Terri L. Braud, HT(ASCP)
HNL Laboratories for Holy Redeemer Hospital
1648 Huntingdon Pike
Meadowbrook, PA 19046
Ph: 215-938-3689
Fax: 215-938-3874

6. Re: release of body parts (E. Wayne Johnson)
Message: 6
Date: Thu, 19 Aug 2021 23:32:13 +0800
From: "E. Wayne Johnson" 
Subject: Re: [Histonet] release of body parts
I will take the other side of this argument.

If you go to the Dentist and he extracts a tooth, it is the usual procedure 
that he gives it to you.
After all it is "your tooth".

Like wise, it's your gall bladder.? The legal department should understand that 
it is your personal property and the mining of it from your body gives the 
hospital no particular right to take control of it any more than they have the 
right to take control of a birthed infant.

E. Wayne Johnson DVM
Enable AgTech
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Re: [Histonet] release of body parts

[E. Wayne Johnson via Histonet]

I will take the other side of this argument.

If you go to the Dentist and he extracts a tooth, it is the usual procedure 
that he gives it to you.
After all it is "your tooth".

Like wise, it's your gall bladder.  The legal department should understand that 
it is your
personal property and the mining of it from your body gives the hospital no 
particular right to take control of it
any more than they have the right to take control of a birthed infant.

E. Wayne Johnson DVM
Enable AgTech
Beijing


John Garratt via Histonet wrote:

I suggest that path labs start a discussion with Risk Management team and 
lawyers to get advice on the tease of tissues to patients.
The uterus in the landfill or the gallbladder at school “show and tell” will be 
sure to get your legal department on edge and the lab’s name in the local paper.
When everybody is stretched to the limit to provide pathology should you also 
be providing a souvenir service when there is a perfectly good gift shop in the 
hospital?
Having a process in place, like using a funeral home with a lab fee attached 
tends to sort out those who just want something to shows their pals at the 
coffee shop.

John

John

Sent from ProtonMail for iOS

On Wed, Aug 18, 2021 at 4:44 PM, Jay Lundgren via Histonet 
 wrote:


It's all fun and games until someone finds a uterus in a landfill.
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Re: [Histonet] release of body parts

[E. Wayne Johnson via Histonet]

I wanted my femoral heads after a dual hip replacement.
I was refused as it was against local policy but the surgeon
was kind enough to take some pictures for me and sawed through one sagitally
so that i could see the pathology.

E. Wayne Johnson DVM
Enable AgTech
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Cartun, Richard via Histonet wrote:

We no longer release any tissue to a patient that comes to our Pathology Laboratory in 
formalin, and our Legal Department supports this decision.  I know that our "Labor & 
Delivery" Unit has released placentas to patients; however, if the specimen doesn't come 
to Pathology, we don't get involved.  For certain types of specimens received in formalin 
(POC, fetus, amputation, etc.) a patient can request the release of their specimen, but they 
have to make arrangements with a funeral home or mortuary to take procession of the specimen 
here at the hospital and, yes, they (funeral home/mortuary) must sign the release form.

Please keep in mind that each state may have statues on the release of human 
tissue to patients or their families.  I also know that some towns here in CT 
have ordinances preventing residents from burying human tissue on their 
property.

Richard

Richard W. Cartun, MS, PhD
Director, Histology & The Martin M. Berman, MD
   Immunopathology/Morphologic Proteomics Laboratory
Assistant Director, Anatomic Pathology
Department of Pathology & Laboratory Medicine
Hartford Hospital
80 Seymour Street
Hartford, CT  06102
(860) 972-1596 Office
(860) 545-2204 (Fax)


-Original Message-
From: Nancy Schmitt via Histonet [mailto:histonet@lists.utsouthwestern.edu]
Sent: Wednesday, August 18, 2021 12:33 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] release of body parts

This email is from outside HHC. BE CAREFUL when opening attachments or links 
from unknown senders.

Hello-
We are seeing a bit more of patients that are requesting to take their body 
parts with them (uterus, POC, etc); I am talking home  - not the funeral home.
Are you using a release of body parts form to fill out with the patient?
Are you draining off the formalin, or sending in formalin with parafilm around 
the lid?
Thank you for your thoughts,
Nancy Schmitt MLT, HT(ASCP)
Pathology Support Services
Dubuque, IA  52001


Confidentiality Notice:
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Re: [Histonet] Dragon

[E. Wayne Johnson via Histonet]

I use Nuance Dragon for making video scripts, transcribing audio stripped out 
of video recordings, and for dictation of documents.
Until one gets it trained it will create very humourous mondegreens of 
technical terms.
But you can add terms like thromboembolic meningoencephalitis and 
extramedullary haematopoesis which it handles with glee.
It is very fast and it can go as fast as I can talk.
I have a very fine Shure desktop microphone and a Jabra headset.  The Jabra 
headset sounds awful on audio recordings
but is great for dictations.  The Shure microphone sounds awesome but doesnt 
match the Jabra headset for dictation speed and
accuracy.  The worst thing about Dragon is that it is Windows-based and they 
dont have a Linux version and it doesnt run under Wine,
so I have to reboot my machine and start windows when i want to use it.

mModal is a 3M product.  3M medical is pretty stodgy and unfriendly and hard to 
get along with.

E. Wayne Johnson, DVM
Enable AgTech
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Terri Braud via Histonet wrote:

We were using mModal and are in transition to Voicebrook which uses Dragon as a 
speech recognition set up with SQ CoPath for both grossing and microscopic 
dictations. We have mixed user feelings, too.  Our PA who does the majority of 
the gross, uses it and has lots of templates preset to simplify the gross. She 
uses voice commands to navigate through CoPath.  She is very fast and seldom 
has an error.  With the pathologists, it's a bit of a mixed bag, somewhat 
dependent on how they embrace the technology change.  Proper installation and 
set up, complete with templates is key.  The use of a headset with mic seems to 
produce the best results as opposed to a gooseneck microphone.  I hope this 
helps.  Terri

Terri L. Braud, HT(ASCP)
Anatomic Pathology Supervisor
HNL Labs, Holy Redeemer Hospital
1648 Huntingdon Pike
Meadowbrook, PA 19046
ph: 215-938-3689
fax: 215-938-3874
Care, Comfort, and Heal

Message: 5
Date: Tue, 9 Feb 2021 16:48:30 -0500
From: Amanda Coscetti 
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Dragon
Message-ID: <3d4a88a7-181e-47b2-b830-64437cae1...@gmail.com>
Content-Type: text/plain; charset=us-ascii

Does anyone use Dragon while grossing? We are looking at switching to Dragon 
and wanted some feedback for use in pathology.
Currently, we use M*Modal for dictations and then have transcriptionists type 
all of the information in Cerner reports.
Thanks!!
Amanda
--
Message: 6
Date: Tue, 9 Feb 2021 17:01:54 -0500
From: Cristi Rigazio 
To: Amanda Coscetti 
Cc: histonet@lists.utsouthwestern.edu
Subject: Re: [Histonet] Dragon
Message-ID: <232b1a9f-8471-48f6-ae43-67b05f97c...@gmail.com>
Content-Type: text/plain; charset=utf-8

I used it exclusively in GI lab for years and loved it but we only did small 
biopsies, so it was super simple.  We use it now throughout the system I work 
in and there are a variety of opinions.  Some of the PAs love it and some 
really struggle.  Same with the pathologists.  So we live in dual worlds with 
both Dragon and MModal.  If you would like to contact me separately I am happy 
to discuss! Thanks Cristi



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Re: [Histonet] Maximum and Minimum Temperatures on Embedding Center Paraffin Tanks

[E. Wayne Johnson via Histonet]

You could warm the paraffin in an oven overnight
to melt it so that the heat of fusion is not extracted
from the embedding center paraffin tanks when it is added
as melted paraffin at 58-65 degrees.

Pairan, Kelly via Histonet wrote:

Good Afternoon,
Recently we starting taking the daily maximum and minimum temperature ranges 
for our embedding center paraffin tanks.  The melting point of our wax is 56oC 
and our current temperature range is 58 to 65oC.  The problem we are having is 
that when we add wax, the temp dips below the range until it melts.  If we turn 
it up, it exceeds the max range.  Any suggestions?

Thanks,
Kelly
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Re: [Histonet] Blades

[E. Wayne Johnson via Histonet]

In China we can buy Leica, Feather, or domestic.
The domestic ones are not uniformly good or bad.
I am really really tired of Feather and I consider that my time 
is worth something so I insist that we use Leica although they
are as "high as a cat's back".

>  ---Original Message---
>  From: raestask via Histonet 
>  To: Histonet@lists.utsouthwestern.edu 
>  Subject: [Histonet] Blades
>  Sent: Nov 18 '20 02:38
>  
>  What breand of blades do people prefer??Rae Staskiewicz UnityPoint Health 
> Methodist PeoriaSent from my Verizon, Samsung Galaxy smartphone
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Re: [Histonet] Tissue processor errors, failures and what to do

[E. Wayne Johnson via Histonet]

Automation is a wonderful thing but it is only a replacement for what people 
used to do by hand.
We have incubators that can be set to 60C and we have a Rube Goldberg-ized 
microwave oven with
a thermal controller and relays (and the not-to-be-forgotten flyback diode) and
a K-type thermal probe covered with aluminum foil (dont try this at home, kids)
so we are ready for any incident or mishap with our cantankerous VIP5.

There is no such thing as a convenient time for it to quit but lots of times 
the trouble
is that it was time to change the solutions and nobody did it even though
anybody could have done it.

If there is nothing we can do to get the VIP5 going again pronto
we finish the processing by hand.
there are repair manuals for some of those machines available online
or you can call the repairman.  If you are out in Bufooee somewhere the repair
manual and the secret repair modes on the machine could be the difference 
between joy
and the slough of despond.

E. Wayne Johnson DVM
Enable AgTech
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Patpxs via Histonet wrote:

Hi Garrey,

The answer is “it depends”.   What you do when a processor fails depends on the 
failure point.  If the tissue is still in dehydrant it gets treated differently 
than if it fails in the intermediate solvent.

Paula

Sent from my iPhone


On Jul 4, 2020, at 10:08 AM, Garrey Faller via Histonet 
 wrote:

Happy 4th to all.
Does anyone have a procedure on what to do when a tissue processor fails or 
alarms.  I want to learn more about the science behind tissue processing so I 
know what to do when the machine fails. This happened to a friend recently and 
I want to prevent my tissues/biopsies from being ruined.
Thanks.
Garrey

Sent from my iPhone
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Re: [Histonet] Formalin fixation for COVID-19 positive tissues .....

[E. Wayne Johnson via Histonet]

Where aldehydes are used for disinfection we consider 30 minutes contact time
for difficult viruses like African Swine Fever (ASF) virus to be adequate.  
Coronaviruses
like PEDv TGE IBV and the SARS virus are much easier to inactivate than ASF 
virus.
I'd consider the SARS virus to be similar to Infectious Bronchitis Virus which 
is systemic
and affects a wide variety of tissues.

Inactivating a virus in a chunk of tissue is more challenging than disinfection 
of contaminated
fomites but I see no reason to think that routine fixation times would not 
completely inactivate
the SARS coronavirus.  The concentrations of aldehyde for fixation are at least 
10 times higher
than the highest concentrations used when aldehydes are employed for 
disinfection.

I'd be more concerned about the container than the contents.

For necropsy samples from your human specimens there is not generally any rush 
to get the results
so the CDC's C.Y.A. time doesn't cause trouble but waiting 72 hours for a 
surgical path result
would seem to be just wasting time.

E. Wayne Johnson DVM
Enable AgTech
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Tony Henwood (SCHN) via Histonet wrote:

Hi Richard,

It will depend on the size of the tissue and the source.
Lung tissue is the major concern. Other tissues not affected as much (based on 
the burgeoning literature on Covid-19).
Routine fixation time are applicable, remembering that the alcohols and heated 
wax will also inactivate the virus (triple whammy).
I suppose that I better add a reference:

https://www.tandfonline.com/doi/full/10.1080/01478885.2020.1734718


-Original Message-
From: Cartun, Richard via Histonet [mailto:histonet@lists.utsouthwestern.edu]
Sent: Tuesday, 21 April 2020 6:51 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Formalin fixation for COVID-19 positive tissues .

How long are you fixing surgical tissue specimens from COVID-19 positive patient's before 
tissue processing?  I know that the CDC is recommending "72" hours for autopsy 
tissues, but, to me, that seems excessive for surgical pathology specimens.  Any 
information that you can share on this subject would be appreciated.  Thank you and stay 
safe.

Richard

Richard W. Cartun, MS, PhD
Director, Histology & The Martin M. Berman, MD Immunopathology/Morphologic 
Proteomics Laboratory Director, Biospecimen Collection Programs Assistant Director, 
Anatomic Pathology Hartford Hospital
80 Seymour Street
Hartford, CT  06102
(860) 972-1596 (Office)
(860) 545-2204 (Fax)
richard.car...@hhchealth.org<mailto:richard.car...@hhchealth.org>


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Re: [Histonet] FW: Microtome at home

[E. Wayne Johnson via Histonet]

"Taking control of the situation is key."

It's very interesting to me that most of the responses have to do with
institutionalized bureaucratic ideas (safety, insurance, liability, regulations)
rather than how to get things done.

I was thinking about how Virchow and Henle and other pioneers would
have been able to get anything done in the face of such reverence
for the stultifying bureaucracy that is the enemy of effectiveness.

Afraid to decide, no action is taken.

/"Auftragstaktik/ can be seen as a doctrine within which formal rules can be selectively 
suspended in order to overcome "friction"."


E. Wayne Johnson DVM
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Mark Tarango via Histonet wrote:

I had heard that CLIA was relaxing things and is not requiring a new # to
work from home right now.  Best to check on the regulatory but FFPE isn't
typically infectious.  The ideal spot would in the garage and not the
kitchen though.

On Thu, Apr 16, 2020 at 3:47 PM Roxana Robinson via Histonet <
histonet@lists.utsouthwestern.edu> wrote:


I do not agree with  this in our current situation or actually any
situation.
There are quidelines in place with CLIA, OHSA  and CAP for protecting not
only the patient but also the employee.  Whether research or not.


Roxana Robinson


On Apr 16, 2020, at 4:58 PM, Patsy Ruegg via Histonet <

histonet@lists.utsouthwestern.edu> wrote:

I agree with this point and as far as clocking in and out, I would

think you could work out something like getting paid piece mill, perhaps
charge per slide or block cut, that way you could do it on your own time
and not have to clock in.


Patsy Ruegg, HT(ASCP)QIHC
Ruegg IHC Consulting
40864 E Arkansas Ave
Bennett, CO 80102
H 303-644-4538
C 720-281-5406
prueg...@hotmail.com



From: Joseph Saby 
Sent: Thursday, April 16, 2020 8:03 AM
To: Porter, Amy ; Porter, Amy via Histonet <

histonet@lists.utsouthwestern.edu>; histonet@lists.utsouthwestern.edu <
histonet@lists.utsouthwestern.edu>; Steven Crochiere 
Subject: Re: [Histonet] FW: Microtome at home


You will need to make sure all pertinent SOPs and EOPs are followed, as

well as all safety guidelines/protocols. Just because it is not human
tissue doesn't mean that it can't have its share of nasties.

Joe Saby

Sent from Yahoo Mail on Android

On Thu, Apr 16, 2020 at 8:21 AM, Porter, Amy via Histonet<

histonet@lists.utsouthwestern.edu> wrote:   Make sure of insurance
coverage and safety for the employee and that they are covered in case of
injury - are they still clocking in and out in some fashion. just
thinking in a bigger box.


From: Steven Crochiere via Histonet 
Sent: Thursday, April 16, 2020 6:36 AM
To: histonet@lists.utsouthwestern.edu 
CLIA would need to inspect the set up in the person home. The same goes for
our pathologists who read slide at home.

Steve

-Original Message-
From: raestask via Histonet [mailto:histonet@lists.utsouthwestern.edu]
Sent: Wednesday, April 15, 2020 7:51 PM
To: Jamie Watson ;

Histonet@lists.utsouthwestern.edu

Subject: Re: [Histonet] Microtome at home

I wouldn't think there would be any problem.Rae Staskiewicz HT(ASCP)Sent

from my Verizon, Samsung Galaxy smartphone

 Original message From: Jamie Watson via Histonet <

histonet@lists.utsouthwestern.edu> Date: 4/15/20  6:44 PM  (GMT-06:00)
To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] Microtome at
home Hello all,Our pathologist has come up with the idea of sending a
microtome and waterbath home to someone that cannot come to work due to
COVID 19.  We are a research lab and work with mouse and rat tissue.  Does
anyone know of any issues with doing this?  I have never heard of anyone
cutting slides at home other than someone with a private business.Thank
you.Jamie___Histonet mailing
listHistonet@lists.utsouthwestern.eduhttp://
lists.utsouthwestern.edu/mailman/listinfo/histonet

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Re: [Histonet] On-line references

[E. Wayne Johnson via Histonet]

And I appreciate Bryan Llewellyn and the other old experienced hands
and I really like Gray and Humason and even Lillie, and the others and the
way they wrote and the way they thought,
and their delight and fascination with the world they were discovering.

E. Wayne Johnson
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Пешков Максим via Histonet wrote:

Many old books are here as e-versions: www.archive.org
--
Maxim Peshkov
Russia
Taganrog
  
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Re: [Histonet] UV LIGHT BOX to kill BLOODBORNE PATH

[E. Wayne Johnson via Histonet]

I really liked that idea of having a secondary copy of the paperwork to 
take to

any potentially dirty area.

I am not very excited about UV light as a disinfectant for paper.
If the paper is dirty, it's dirty.
Incineration is a great disinfectant method.
Fire is another good one for paper (as a substitute for incineration).

E. Wayne Johnson DVM
Enable AgTech
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Paula via Histonet wrote:

Hello,

  


Has anyone ventured out and looked into a UV light instrument, like a box
that can kill any bloodborne pathogens that are on paper (grossing
area..where the patient req form is used)?

  


The administrator has a grand idea about placing any paperwork that contains
smudges/smears inside a UV lightbox to kill potential pathogens.

  


Thank you in advance,

Paula

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Re: [Histonet] Tissue Tek TEC 6 Embedding Centre

[E. Wayne Johnson via Histonet]

I am skeptical that parts are not available for these relatively new 
(fifteen years old) machines...


Of course reliability is a serious concern in any case.

E. Wayne Johnson DVM
Enable AgTech
Beijing

Etheridge, Sandra AGRI:EX via Histonet wrote:

Hi everyone,

We are looking to purchase a new embedding centre and was wondering if anyone 
has the new Tissue Tek TEC 6?  Our current Tissue Tek TEC is 15 years old and 
parts are no longer available for repair.

We are comparing the TEC 6 to the Leica HistoCore Arcadia system.  Just 
wondering if anyone can give some feedback with regard to pro and cons of 
either unit.
Much appreciated!

Sandra Etheridge

BC Ministry of Agriculture
Plant and Animal Health Centre
Histology/IHC
1767 Angus Campbell Road
Abbotsford, BC V3G 2M3
# (604) 556-3120

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Re: [Histonet] Processing Veterinary Samples

[e wayne johnson via Histonet]

The vet samples can be processed just any other animal tissue whether they be 
human or rat. People is animals too. Formalin fixed there are no special safety 
issues. E. Wayne Johnson DVM Enable AgTech Beijing ewj Email:e...@pigs.ag 
Signature is customized by Netease Mail Master On 04/16/2019 08:05, Miranda 
Giorgi via Histonet wrote: Hello, Our AP lab was recently approached by a 
Veterinary Pathologist to process, embed, and cut H slides of their 
postmortem animal samples.  If we consider taking on this work are there any 
concerns with variation in processing or safety protocols we might need to be 
aware of with regards to veterinary samples compared to human samples? Any 
information or experience you can share would be appreciated.  Thank you! 
Miranda Giorgi, HTL (ASCP)cm Histology Manager Incyte Diagnostics 509-892-2744 
This e-mail and any attachments may contain CONFIDENTIAL information, including 
PROTECTED HEALTH INFORMATION. If you are not the intended recipient, any use or 
disclosure of this information is STRICTLY PROHIBITED; you are requested to 
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notify the InCyte Privacy Officer at priv...@incdx.com or call (509) 892-2700. 
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Re: [Histonet] PIG processing tissue

[E. Wayne johnson via Histonet]

We routinely examine pig heart as part of diagnostic histopathology

and generally we are looking for the lesions of Vitamin E / Selenium 
deficiency.


Vitamin E and organic Se are expensive and often deficient in commercial 
piglet diets.


We simply fix with a formalin based fixative and process routinely to 
paraffin


and stain with our H & E.

-

E. Wayne Johnson

Enable AgTech, Beijing.

"Never do anything mean to a pig.  They'll squeal on you every time."




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Re: [Histonet] Eosin

[E . Wayne Johnson 朱稳森博士 via Histonet]

You might try some combination of eosin and biebrich scarlet and/or 
phloxine.


Adding a little acetic acid might help.

http://www.stainsfile.info/StainsFile/downloads/eosin.pdf

I am quite fond of Biebrich Scarlet and like way it stains brain and 
heart particularly.


Biebrich is much more red than eosin.

Phloxine is an aggressive pink.

*

A bit punny about the fish looking washed out.

*

"There was something fishy about the butler.

Probably a Pisces working for scale." - Phil Proctor



On 08/30/2016 10:25 PM, Cameron, Elizabeth wrote:

Hi,
I have been staining fish tissues fixed in Davidsons with H, and the 
researcher would like the eosin to be more intense.  Our standard protocol works 
well for our own tissue, but the fish look much more washed out.  I am using 
alcoholic eosin Y, have tried both water and alcohol before and I have varied the 
alcohol differentiation steps after the Eosin.  I also extended the time in Eosin 
and increased the wash after bluing to make sure the sections are not basic.  Any 
suggestions would be appreciated.
Thank you.

Elizabeth M. Cameron, HT(ASCP), QIHCCM
Lead Histologist
Mid Coast Hospital
123 Medical Center Drive
Brunswick, ME 04011
(207) 373-6573




--
E. Wayne Johnson 朱稳森博士
Enable AgTech Consulting
恩睿康农业技术咨询有限公司
Beijing
188 1088 3205

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Re: [Histonet] Start up Vet/Animal histology laboratory

[E . Wayne Johnson 朱稳森博士 via Histonet]

We have our own lab in Beijing and we do both of those things.

We have a consulting veterinary practice and the main purpose of the lab 
is to support that veterinary practice.  It would be unusual in the USA 
to have histopath in a vet lab but in China there was no diagnostic 
laboratory that did any histopathology at all.  We have some cooperation 
with a university and we get some research samples from them.


We had some difficulty procuring reagents at first.

Fixed samples from the field or from researchers are often in horrible 
shape.


It really is a disaster when samples from expensive and difficult to set 
up research are ruined by inept collection and poor fixation...


We are primarily interested in pigs but we get research samples from 
mice and chickens sometimes.


If there is a good liaison with the researchers it works well.

I have a team of people who know how to select and necropsy pigs for 
diagnosis and they can do a good job.  We made a video to show people in 
the field how to collect and manage diagnostic samples.


I knew that personnel was difficult.  If you hire and train young men, 
they will leave and go work for someone else after they think they know 
what they are doing.  If hire young women, they will get married and 
have babies and can cause many problems.  I am tired of hiring people 
with advanced degrees (MS, PhD).  They tend to be lazy and are rather 
hard to train.  Bright young folk who have the willingness to learn are 
a joy.  The surprising thing about a MS or PhD is what it ain't.


In the future I am considering hiring taxi cab drivers for everything. 
They are smart.  They are ambitious.  They know how to take directions. 
They are self-starters and independent workers.


On 07/10/2016 07:53 AM, Linda wrote:

  Hello Histoland,
I am inquiring if anyone has started their own lab to do histology services for 
veterinarians or animal research tissues?
Or anyone who has started a private lab?
What are your experiences?  What didn't you expect?
Thank you in advance.
Regards,
Linda Dee, BGS,HT(ASCP) lmd...@yahoo.com





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Re: [Histonet] Pap stain without xylene

[E. Wayne Johnson via Histonet]

Xylene is becoming more and more of a nuisance material and a problem 
for us in use and in disposal.  We are still able to use it but with 
increasing concern.


We have been able to eliminate xylene from our staining procedures 
altogether in our small laboratory.


We use a hair dryer to dry the slides in the rack after water or alcohol.

Since we read our slides right away, we have been using cedarwood oil to 
clear and mount the slides.  It's cheap, and makes lovely slides with no 
crystals or "floaties" or other artifacts.  It's easy to clean up 
without any xylene or toluene. It's not permanent but we can remount the 
slides with a permanent mounting medium if we need to keep the slides 
for some reason.


*

We also have found several different methods for eliminating xylene from 
the paraffin infiltration process,


and we have not used xylene for dewaxing for more than 2 years now.


On 05/26/2016 03:07 AM, Mike Toole wrote:

Thank you Beth,

It’s good to know that someone else had tried this and had very good results. 
The method you suggest is very much in line with the recommendations from René. 
He did recommend using a drying oven at 60°C to help ensure complete removal of 
any water or alcohol. And, that absolute dryness was a requirement for 
coverslipping without artifact such as the appearance of sand like grains or 
cornflakes.

Just to reiterate, was the field method performed at ambient temperature 
without the aid of a drying oven? And, just a thought, I suppose if the lack of 
electricity was an issue in a field setting, that perhaps a solar oven made 
with plywood and glass could be used to elevate temperature for drying.

Do you  know if altering the method for final clearing would require validation?

Mike


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Re: [Histonet] picric acid

[E. Wayne Johnson via Histonet]

I read "Chemical Magic" and the "Anarchist's Handbook" in high school 
many years ago.  That was back when boys carried pocket knives and 
sometime took their shotguns to school to show their friends.  I still 
occasionally make a little bit of NI_3 for fun.


We've tried to make picric acid explode and have taken it out in the 
yard and burned it and beat on it with hammers. It's really not all that 
easy to make it go "BOOM" or even make crisp pops. Just don't mix it 
with heavy metals.


I agree strongly that it should be handled with respect, and with gloved 
hands like any staining powder.  Store it wet, but the /~~paranoia~/~ 
about picric acid is exactly that.


And on the part of the cops and bomb squads who take picric acid out and 
detonate it amid much drama and fanfare, heavy on the delusions and 
illusions of grandeur.


E. Wayne Johnson, DVM

Enable Ag-Tech

Beijing

On 05/07/2016 12:01 AM, Julio Benavides wrote:

Thank you so much everybody for your help!!



"Morken, Timothy" <timothy.mor...@ucsf.edu> escribió:


Here's another good document on how to handle picric acid powder

www.ehs.wisc.edu/chem/SafeHandlingOfPicricAcid.pdf



-Original Message-
From: Morken, Timothy via Histonet 
[mailto:histonet@lists.utsouthwestern.edu]

Sent: Friday, May 06, 2016 8:28 AM
To: Julio Benavides
Cc: Histonet
Subject: Re: [Histonet] picric acid

Julio, you can just pour water into the container. We always 
oversaturated so that a layer of water was on top of the powder.


Look at this explanation
http://oag.ca.gov/sites/all/files/agweb/pdfs/cci/safety/picric.pdf

or read the text below if you cannot open this.  This contains 
instructions on how to properly store picric acid powder, and how to 
deal with dry powder found in the lab.


Long ago I had the pleasure of discovering a batch of six 2kg bottles 
of dry picric acid in our "bunker" where we stored flammables. Over 
10 years old according to dates on the box. We called in the fire 
department to take care of it. They hosed it down, removed it and 
disposed of it; how, I don't know.



Tim Morken
Pathology Site Manager, Parnassus
Supervisor, Electron Microscopy/Neuromuscular Special Studies 
Department of Pathology UC San Francisco Medical Center



+++
PICRIC ACID HAZARDS
Mark Cameron, CIH
Every couple months, an article appears in the local paper about a 
bomb disposal team removing picric acid that was found in a 
laboratory. The material is usually taken to be blown up. So why is 
picric acid considered so dangerous? Well, let’s look at the history 
of the use of Picric Acid and see what can be done to avoid those 
types of situations.


Picric Acid (2,4,6 Trinitrophenol) is frequently found in forensic 
laboratories for use in the Christmas Tree stain (1) and for Urine 
detection (2). Histology uses include connective tissue stain 
(Jullien’s picroindogocarmine and Van Gieson’s picro-acid fuchsin), 
cytoplasmic stain (Van Gieson’s with iron hematoxylin), woody 
sections (picro aniline blue) and as a fixative agent (3). It was 
used in medicinal formulations in the treatment of malaria, 
trichinosis, herpes, smallpox and antiseptics. A one- percent 
solution was also used in the treatment of burns (4).
British Chemist Peter Woulfe discovered picric acid in 1771. Picric 
acid was named from the Greek word pikros, which means “bitter” due 
to its bitter taste (5). It was used to dye silk and wool yellow. 
Workers making picric acid during World War I were called “canaries” 
because their skin was stained yellow (6).


The explosive characteristics of Picric acid were discovered early. 
In 1885, experiments with picric acid were conducted in Lydd, England 
and the English adopted it as an explosive material called Lyddite in 
1888. It was used extensively in bombs and grenades during World War 
I (7). Anhydrous Picric acid is similar to TNT. It needs usually 
needs a “booster” such as a primer to create the explosion. However, 
as a strong acid, picric acid attacks common metals (except tin and 
aluminum) creating explosive salts, which are shock-sensitive. Bombs, 
mines and grenades were coated with tin or ashpatim to prevent the 
picric acid from contacting the metallic shell (8).


Several catastrophic events involving picric acid have occurred. On 
December 6, 1917, an ammunition ship in Nova Scotia carrying 2,300 
tons of picric acid as well as 400,000 pounds of TNT caught fire and 
exploded. Over 1,900 people were killed immediately and 9,000 were 
injured (9). Shock-sensitive metal picrates demonstrated their 
hazardous nature on May 1, 1916 when a fire at a French ammunition 
factory caused molten picric acid to flow onto the concrete floor. 
Calcium picrate was formed and detonated, killing 170 people (10).

06/18/02

Have there been any explosions in laboratories? There are no 
documented instances of spontaneous detonation of picric acid in a 
laboratory (11). 

Re: [Histonet] RE: Glassware Cleaning again

[E. Wayne Johnson]

Bromcresol means Bromocresol.

Faint not, but I certainly can't say you won't dye if you leave out the O.

The o is subject to elision  due to its difficulty in pronounciation.

Such an occurrence of elision between 2 consonants is called syncope.

In English writing an elided vowel is often replaced with an apostrophe to
indicate the elision and perhaps demonstrate the dialect of the speaker.

Colloquial dialects in writing are too informal for stuffy scientific 
and medical types, and indeed

special meaning is liable to be construed to the presence of a spurious
punctuation in the name of a chemical.

We don't see {brom'cresol} in
the lab {purple, green} but we can hear of an existence
where it lives happily under other nomenclature.

So bromcresol means bromocresol.

And as in American politics,
the removal of an O might seem to make a difference visually
and even might be comfortable to some,
but actually would amount absolutely nothing in terms of what is
being represented, or the dying that is going on.



On 8/16/2014 12:35 AM, Cooper, Brian wrote:

I noticed the discrepancy in spelling too.  I looked online for like 30 minutes, and 
couldn't find anything called Bromcresol.  Found a lot of vendors selling 
Bromocresol Purple (and Green for that matter).  Best I can figure is that this is a typo 
on CAP's checklist (that's been there for several years now).

Thanks,

Brian D. Cooper, HT (ASCP)CM | Histology Supervisor
Department of Pathology and Laboratory Medicine
Children's Hospital Los Angeles
4650 Sunset Blvd MS#43- Los Angeles, CA 90027
bcoo...@chla.usc.edu

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Laurie Colbert
Sent: Friday, August 15, 2014 9:13 AM
To: Histonet Post (histonet@lists.utsouthwestern.edu)
Subject: [Histonet] Glassware Cleaning again

Is bromocresol purple the same as bromcresol purple?  The CAP question 
regarding glassware cleaning refers to bromcresol purple, but I ordered a 
powder and it is labeled as bromocresol purple.

Laurie Colbert, HT (ASCP)
Histology Supervisor
PATH MD
8158 Beverly Blvd.
Los Angeles, CA  90048
(323) 648-3214 direct
(424) 245-7284 main lab

The information contained in this transmission may contain privileged and 
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review, dissemination, distribution, or duplication of this communication is 
strictly prohibited. If you are not the intended recipient, please contact the 
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Re: [Histonet] specimen marking ink

[E. Wayne Johnson 朱稳森]

Tattooing is not only for dissidents, miscreants, the wayward,
and the Llewellyn-ites among us.

Sheep, rabbits, pigs, cattle, horses, stoats, and goats can be tattooed 
also,
not so much of an expression of individuality as the need for permanent 
identification.


http://www.enasco.com/c/farmandranch/Livestock%20Identification/Tattooing/


On 3:59 AM, Davis, Cassie wrote:

Hi Histo World, as I was cutting to day I was thinking why don't we see if we could get 
specimen marking ink directly from a tattoo vendor? When I first started in histo I was told 
the ink we use was actually tattoo ink. As we know as soon as somebody labels something as a 
medical supply the price is increased. Just a cost saving thought, I mentioned it 
to my immediate supervisor but she think it would be a liability issue. I thought we could 
test/validate it on skin tissue left over from a mastectomy or extremity. Any thoughts?

Cassandra Davis
cda...@che-east.org
302-575-8095



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Re: RE: [Histonet] Gross lab seniors

[E. Wayne Johnson 朱稳森]

Face velocity is simply the airflow rate in CFM divided by the area of 
the hood opening in square feet.


A smaller opening at the same flow rate gives a higher face velocity.

Titanium tetrachloride in a small plastic squeeze bottle can be used to 
generate smoke.



On 3:59 AM, WILLIAM DESALVO wrote:

We use a company called C-Scan Technologies, Phoenix, AZ. The way they test all 
our gross dissection stations is by testing for directional or smoke containment 
and face velocity. We also check th they external pathway is clear and if the unit 
has a filtering system, the filters are changed regularly. The air flow 
measurement is Feet per minute (FPM) for face velocity and includes width, height, 
depth and total square ft for the working area. They exhaust flow in CFM. Face 
velocity minimum requirement is 100 fpm, exhaust flow requirement is500 cfm. 
Face velocity fluctuates depending on the room and the air exchange rate for the 
area. I have always felt the face velocity is most important to gross dissection 
personnel. There needs to be adequate draw away from the employee, no matter the 
physical conditions of the room.

William DeSalvo, BS HTL(ASCP)
Production Manager-Anatomic Pathology
Chair, NSH Quality Management Committee
Owner/Consultant, Collaborative Advantage Consulting


   

From: vickroy@mhsil.com
To: histonet@lists.utsouthwestern.edu
Date: Fri, 31 Jan 2014 12:46:08 -0600
Subject: [Histonet] Gross lab seniors


We have several gross lab senior grossing stations that are vented outside.  
Our engineering asked today whether the airflow should be checked yearly like 
other exhaust hoods.   Problem is there is not a door like other hoods of 
course and how would you measure the airflow?   Recommended airflow is 500cfm 
however clearly the airflow at the working surface is not anything close to 
that.   I wondered how anybody else monitors the gross lab seniors or do they 
at all.   CAP used to ask about documentation for checking hoods however I 
can't recall them ever checking on grossing stations.  We change filters 
annually  only since they are vented outside.

Jim

James Vickroy BS, HT(ASCP)

Surgical  and Autopsy Pathology Technical Supervisor
Memorial Medical Center
217-788-4046



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Re: [Histonet] In situ PCR

[E. Wayne Johnson 朱稳森]

We don't do in-situ PCR, but the principle is that with formalin-fixed 
tissues

your amplified product is trapped in the protein matrix on the slide.




On 3:59, Sarah Dysart wrote:

Anyone out there do this?  If so, during the PCR step you are amplifying your 
gene of interest, where does the amplified product go?  Each step of the PCR 
(from how I am understanding this...I'm new to molecular biology protocols...) 
separates the double stranded sequence then copies it, and this goes on and on 
for 30-40 cycles...where does the product go?  Does it just wash off?  If not 
how is it binding to the tissue?

Sarah Goebel-Dysart, BA, HT(ASCP), QIHC (ASCP)
Histotechnologist
Mirna Therapeutics
2150 Woodward Street
Suite 100
Austin, Texas  78744
(512)901-0900 ext. 6912


   



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Re: RE: [Histonet] (no subject)

[E. Wayne Johnson 朱稳森]

We work in the veterinary medical field and 98% of our work is with 
swine diseases
including some feed additive safety evaluations.  We provide pictures of 
all the lesions
that we see as part of the report we provide to farms.  It definitely 
takes more of our
time than it would to give a pathomorphologic description in English and 
Chinese
but we believe in transparency even though it might expose us somehow, 
realizing

that some things in pathology are art.

We are providing service and we think the best way is to be as transparent
as possible in everything that we do.

Stonewalling and secrecy generally breed contempt.
We try to be at our best even when we dont feel like it.

It is true that our subjects have economic value only and they are 
expendable

and the pigs are unlikely to see us in court.  But I think that even in
situations where there are emotions as with pets, and where there are
real human concerns about quality of care for human patients, transparency
and participation of all those concerned in the diagnositic and
evaluative process generally creates good will and cooperation and
good outcomes.


On 3:59, CHRISTIE GOWAN wrote:

Hi Clay,

Most hospitals have a tissue committee that makes decisions about surgical 
specimens. These decisions become hospital policy. Most hospitals state that 
once a specimen is removed it essentially becomes the property of the hospital. 
Release of specimens is usually after it has been accessioned and grossed in 
for pathology reveiw. There is usually a form for that type of release. The 
hospital would be at risk if they let the specimen leave without following 
hospital policy. I am not saying it is right or wrong but I can see the logic 
in it. Once the specimen has been processed through the hospitals pathology 
department then blocks and slides can be requested for 2nd opinions. Good luck 
in your search for answers.

Christie Gowan



To: histonet@lists.utsouthwestern.edu
Date: Thu, 12 Sep 2013 12:38:10 +
From: claymi...@hotmail.com
Subject: [Histonet] (no subject)




I need a little help on a Patient’s rights question.


It is my understanding that when a patient has a procedure, the patient has the 
right to request that those specimens be examined by a laboratory of their 
choosing.  i.e. EGD, colonoscopy, etc


I am in Arkansas.


My father had a procedure yesterday at a local hospital.  I manage a pathology 
laboratory that specializes in the type of tissue that the procedure procured.  
When it was requested that the tissue be sent to my laboratory, the hospital 
staff refused to fulfill the request.  We asked multiple times for a release 
form so my father could take his tissue with him.  The administration employee 
we spoke to said there was no such thing and that patients were not allowed to 
take their specimens. One employee going so far as stating that if we wanted 
the tissue sent somewhere aside from their contracted laboratory, that the 
procedure would be canceled and my father would have to go somewhere else.   My 
father, not wanting to cause a fuss, let the issue go.


Questions:

Is it legal for a hospital to require that tissue specimens be sent to a lab 
they are contracted with?


Are there any other actions we could have taken to make our requests be honored?



This is not meant to insult the hospital, but to give an explanation of the 
situation for context in answering the questions.

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Re: RE: [Histonet] Oil red O

[E. Wayne Johnson 朱稳森]

While you're at it, maybe some one can explain what the
-wah- in diddy-wah-diddy means?



On 3:59, Morken, Timothy wrote:

Just like Formalin, a brand name now used interchangeably with formaldehyde.

And your trivia of the day:

Formaldehyde was the first polyatomic organic molecule detected in the interstellar 
medium

Courtesy of Wikipedia.

Tim Morken
UCSF Pathology

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of WILLIAM DESALVO
Sent: Monday, September 09, 2013 2:00 PM
To: Rene J Buesa; Ada Feldman; histonet
Subject: RE: [Histonet] Oil red O

I believe the answer should be - Oil Red O is a brand name (not uncommon in the 
dye indutry) and represents the two dyes found in the compound, Solvent Red 27 
and Sudan Red 5B

William DeSalvo, BS HTL(ASCP)

   

Date: Mon, 9 Sep 2013 13:32:08 -0700
From: rjbu...@yahoo.com
To: adafeld...@anatechltdusa.com; histonet@lists.utsouthwestern.edu
Subject: Re: [Histonet] Oil red O
CC:

There you have it!
René J.



From: Ada Feldmanadafeld...@anatechltdusa.com
To: histonet@lists.utsouthwestern.edu
Sent: Monday, September 9, 2013 4:13 PM
Subject: Re: [Histonet] Oil red O


As you are finding out the answers to dye nomenclature you can had these endings to 
your list of the O' in Oil red O:
Oil red EGN
Oil red 4B


Ada Feldman
Anatech Ltd.
1020 Harts Lake Road
Battle Creek, MI 49037

Phone: 800.262.8324
Phone: 269.964.6450
Fax: 269.964.8084
adafeld...@anatechltdusa.com




On Sep 9, 2013, at 3:59 PM, Connolly, Brett M wrote:

 

I'm not sure, but whatever you do... don't store it next to the Sudan Black 
B...you'll stink up the lab!!

Brett M. Connolly, Ph.D.
Principal Scientist, Imaging Dept.
Merck  Co., Inc.
PO Box 4, WP-44K
West Point, PA 19486
brett_conno...@merck.com
T- 215-652-2501
F- 215-993-6803






-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of P.E.
Visser
Sent: Monday, September 09, 2013 3:41 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Oil red O

Hi all

I was requested where the O stands for. who has any suggestion.



Regards Piet Visser

Histotech Bronovo The Netherlands





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Re: [Histonet] Bone/cartilage/epithelial tissue stain

[E. Wayne Johnson 朱稳森]

At one time I did a lot of work on cartilage and growth plate
and used Toluidine Blue and Fast Green.  T-blue (buffered appropriately) 
stains the
proteoglycan a lovely metachromatic blue and the bone and most 
everything else
is green.  The nuclei of cells are also green since we used no nuclear 
stain, but

that was not a problem for our work.

The cartilage is in stark contrast to the bone because of the high 
carbohydrate

(glycosaminoglycan/proteoglycan) content of cartilage.

On 3:59, Rui TAHARA wrote:

Hi,
Would anyone suggest me what staining is
best to color differentiate between cartilage and bone and epithelial tissues
in avian embryos?

I have been trying Mallory Trichrome for
embryos but recently I was suggested that Mallory Trichrome stains cartilage 
differently
in embryos compared to adult samples since Aniline blue stains fiber that may
not develop in early embryos. There is some protocol that modified the Mallory 
Trichrome
that could be applied to embryos. However, the resulting colors of all tissues
look all purple-ish and difficult to tell the cartilage from the weak blue
stain from aniline blue.

Currently I am thinking to try out Alcian
blue/Hematoxylin and Eosin stain (Ehrlich’s hematoxylin). The purpose of the
staining is to look at interaction between ossification and epithelial 
development
so I think alcian blue for staining cartilage works but I am wondering if there
is any other staining combination with alcian blue exist for visualizing bone
and epithelial tissue (e,g. alcian blue/alizarine red with other staining?).
   



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[Histonet] PCR on paraffin sections

[E. Wayne Johnson 朱稳森]

Hi.

We would like to start doing PCR on paraffin fixed tissues from pigs..

It has some appeal to us because we can see the lesions in the tissue
and then we can go after the causative agent post-hoc.  It may be useful
in cases where we have clear lesions on histopath but not the right 
fresh tissues

for PCR.  We are very limited on what we can do with IHC because
of the extreme difficulty of procuring the primary Ab.  It's very
simple to make PCR primers here.

We do have PCR up and running for all of the diseases that we
are interested in.  I understand that the process is basically transferring
the sections to a small tube instead of a slide, then dewax with a little
xylene, and remove the xylene with alcohol, then digest the section
to release the viral RNA.

Does anyone have any recommendations, advice, or experience with the 
digestion

step, and can suggest an appropriate enzyme mix for this digestion?

Yes I am considering in-situ hybridization but we have working PCR methods
so that seems to be a simple logical step.

E. Wayne Johnson
Enable Ag Tech
Beijing.

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Re: Re: [Histonet] Blade Rationing

[E. Wayne Johnson 朱稳森]

A stingy person is called an Iron Rooster (tie gong ji) in Chinese.

An iron rooster won't turn loose of even one feather.

The scratches that appear on slides cut with blades that should have 
been changed --we call
them iron rooster tracks.  Sort of a pun since many people think Chinese 
characters

look like chicken tracks.

We don't worry about what the blades cost.  Our clients demand
good answers, and we send pictures of the lesions with our reports,
so we need good results.

On 3:59, Rene J Buesa wrote:

At 200 blocks/day x 5 days/week x 4 weeks/month = 4000 blocks/month which means 
that the 3 of you will have to use 1 blade every 80 blocks including trimming 
and sectioning which is ABSOLUTELY RIDICULOUS!
The cost of blades, especially the better ones, are going up and you can save 
by using one blade to trim and another to make the final section, but at the 
rate your manager wants the quality will be compromised.
The norm (if there is a norm at all) is that a histotech will probably change 
blades every 5 to 10 blocks if the infiltration is good and there are no decals involved 
in the process.
Lets assume that you can hold to 1 blade every 10 blocks, that will mean that 
during 1 month you will use 400 blades = blades boxes.
Find out how many you are actually using now and you will have an idea of your 
present blades usage.
Additionally dull blades not only compromise the quality of the sections but 
also reduce sectioning productivity and what you may be saving in blades are 
going to increase in histotech time and total section production costs.
René J.

From: Teresa Mooretmoor...@gmail.com
To: histonet@lists.utsouthwestern.edu
Sent: Monday, June 17, 2013 5:10 PM
Subject: [Histonet] Blade Rationing


I work in a hospital, there are three of us on this particular shift and we
cut approx. 200 blocks, give or take a few.  Our histo lab manager is
telling us we should only be using one pack of blades (50 per pack) a
month.  I'm wondering what other techs think of this especially lab
managers and supervisors.

tmoor...@gmail.com
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Re: Re: [Histonet] teeth sectioning

[E. Wayne Johnson 朱稳森]

Jack,
That sounds really awesome.

I did some work with the teeth of sows (female pigs) from
specimens collected at slaughter.  Those are very difficult to decalcify,
and when finished, are likely to have no nuclear detail remaining.

Interested to hear what you learn

Wayne Johnson
Beijing Enable Ag Consulting
Yuanmingyuan West Road Meiyuan Com,


On 3:59, Jack Ratliff wrote:

Mes,

This is a very good question and I look forward to answers from individuals 
that have accomplished this with PMMA and a rotary microtome with 
tungsten-carbide knives. If you are talking about an undecalcified specimen 
embedded in PMMA, then I would imagine that the age of the rat could affect the 
ability to achieve an adequate infiltration of the resin. Again, I look forward 
to what others have to say about their success by the method you have outlined.

On the other hand, I know you can achieve the micron thickness you desire if 
you were to use a non-contact femtosecond laser! The machine I am talking about 
is basically a laser microtome manufactured by Rowiak in Germany and it is 
officially called the TissueSurgeon. In fact, Dr. Heiko Richter from Germany 
has accomplished what you ask with human teeth, revealing the full anatomy of 
the tooth and even with ameloblasts on the enamel surface!

I would be interested to hear more about your project. I will be traveling to 
Germany one week from today to work with this laser microtome until the end of 
the month so I could arrange to have laser cut sections made for you if you are 
interested and unable to make your cuts using PMMA and a rotary microtome. If 
you would like more information, please feel free to contact me by email reply.

Best Regards,

Jack



Jack L Ratliff
Owner/Histologist, Ratliff Histology Consultants, LLC
Chairman, Hard Tissue Committee - National Society for Histotechnology



On Mar 1, 2013, at 7:03 PM, mesruh turkekulturke...@gmail.com  wrote:

   

Dear Histonetters,


I have one more question. Is it possible to obtain 5-10um thick sections of
PMMA embedded teeth using regular Leica paraffin microtome (RM2265)
equipped with disposable tungsten carbide blade?

Thanks,
Mes
On Fri, Mar 1, 2013 at 7:40 PM, mesruh turkekulturke...@gmail.com  wrote:

 

Dear Histonetters,


I am studying bone and teeth growth in rat maxilla. I will inject calcein
green and would like to fix, embed and sections the rat maxilla.
Any suggestions for the best method to fix, embed and section the samples
for fluorescnet microscopy?

Thank you very much!

Mes HTL (ASCP)
Memorial Sloan-Kettering Cancer Center

On Fri, Mar 1, 2013 at 1:01 PM,
histonet-requ...@lists.utsouthwestern.edu  wrote:

   

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than Re: Contents of Histonet digest...


Today's Topics:

   1. FNA Clia Guidelines (PicheGrocki, Jessica)
   2. RE: FNA Clia Guidelines (Horn, Hazel V)
   3. GSH Symposium April 12-14 (Zimmerman, Billie)
   4. FSH abstracts deadline (Jerry Santiago, MSEd, HTL (ASCP) QIHC)
   5. (no subject) (Vikrant Piprode)
   6. The GSH 4oth Anniversary Meeting (David Kemler)
   7. QIHC (Renee H. Workman)


--

Message: 1
Date: Thu, 28 Feb 2013 20:26:30 +
From: PicheGrocki, Jessicajpiche-gro...@wtbyhosp.org
Subject: [Histonet] FNA Clia Guidelines
To: histonet@lists.utsouthwestern.edu
histonet@lists.utsouthwestern.edu
Message-ID:

631955447a364b45b9458d2905635110655d2...@win08-mbx-01.wtbyhosp.org
Content-Type: text/plain; charset=s-ascii

Hi All,

Quick questionwhat are the Clia requirements for Fine needle
aspirate procedures? Is it considered high complexity testing? And who
prepares the slides when the needle is handed off?

Thank you,

Jessica Piche, HT(ASCP)



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Message: 2
Date: Thu, 28 

Re: Re: [Histonet] grossing tools

[E. Wayne Johnson 朱稳森]

I intend to make us some of these tools.

It's a great idea.


On 3:59, Rene J Buesa wrote:

How much squeezing? Tissues have certain elasticity and after the squeezing they will bounce 
back as a thicker slice. Free hand sectioning I think is always better.
René J.

From: Weems, Joyce K.joyce.we...@emoryhealthcare.org
To: 'Bill B.'bill...@mindspring.com; 
histonet@lists.utsouthwestern.eduhistonet@lists.utsouthwestern.edu
Sent: Wednesday, February 13, 2013 11:09 AM
Subject: RE: [Histonet] grossing tools

I have found squeezing the tissue (for the block sections) between two 
cassettes held back to back gives the firmness needed to trim good thin 
sections.

Joyce Weems
Pathology Manager
678-843-7376 Phone
678-843-7831 Fax
joyce.we...@emoryhealthcare.org



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5665 Peachtree Dunwoody Road
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-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Bill B.
Sent: Wednesday, February 13, 2013 9:58 AM
To: histonet@lists.utsouthwestern.edu
Subject: Re: [Histonet] grossing tools

At 5:38 PM + 2/12/13, Bruce Gapinski wrote:
   

 I'm sure we are not the only histology lab that deals with thick grossed 
specimens. Has anyone tried the new gross tools by Sakura? Or anything else that can 
cut ONE nickel thick.
 

A bane indeed, not only for histotechs, but for those doing the grossing. Our 
big problem is breast biopsies. The combination of fat and fibrous tissue makes 
it hard (impossible) to get consistent thicknesses manually.

I have been trying to find some kind 'jig' that would hold inked breast 
biopsies of widely varying sizes that could guide the knife as the biopsy is 
bread-loafed to get consistent thicknesses. I'm not sure what words to look for 
or search for.

OTOH, given our difficulty in getting paid for the tech component (we are an 
independent lab, long used to global billing), I dont want to spend a fortune 
for something.

All advice is super-greatly appreciated.

Bill

--
__
Bill Blank, MD
Heartland Lab

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Re: Re: [Histonet] RNA isolation form stained slides

[E. Wayne Johnson 朱稳森]

I read one paper or abstract recently where sections were made at 50 
microns then used
successfully for RT-PCR.  I am pretty sure that it was PRRS virus that 
they were pursuing.


On 3:59, Mark Tarango wrote:

  I would try getting more sections from LCM and extracting into 7.5 uL, if
that volume will be enough to for your PCR reactions.

On Friday, November 2, 2012, Vanessa Orsini wrote:

   

With the kit I'm extracting in 10ulthe problem is that I have too use
few stained cells isolated with the LCM...so even if I increase the number
of slides I'll never have a lot of material...

Inviato da iPhone

Il giorno 2 nov. 2012, alle ore 16:44, Mark Tarango
marktara...@gmail.comjavascript:_e({}, 'cvml',
'marktara...@gmail.com');  ha scritto:

Have you tried using more sections during extraction?  Can you extract
into a smaller volume?

On Friday, November 2, 2012, Vanessa Orsini wrote:

 






Hello,

I need to extract RNA for a RT-PCR after Laser Micro
Dissection on xGal stained slides.

I tried using sections from unfixed frozen organs. I fixed the
sections in EtOH70% for 10min and then I stained them with xGal for 3h at
37°C.
After air drying I cut out with the LCM and extract RNA with the PicoPure
kit
from Applied Biosystem. So far I didn’t manage to get enough RNA.

I tried to add RNase inhibitors to all the solutions but it
didn’t help.



Any idea/suggestion?

Do someone think it would be better to do a LacZ antibody staining
on FFPE sections and extract RNA with an appropriate kit? The RNase would
they
be less active?



Thank in advance for any help you can give me J Vanessa


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Re: [Histonet] Re: special stains for demonstrating intra cytoplasmic DNA viral inclusion bodies

[E. Wayne Johnson 朱稳森]

Rene' ,

What procedure do you recommend for Giemsa staining of tissues?

There seem to be several different variant protocols out there.

Wayne

Enable (Enruikang) AgTech Consulting
Beijing


On 10/14/12 22:56, Rene J Buesa wrote:

Try Giemsa for tissues
René J.



From: anjan kumardrvet_an...@hotmail.com
To: histonet@lists.utsouthwestern.edu
Sent: Saturday, October 13, 2012 2:48 PM
Subject: [Histonet] Re: special stains for demonstrating intra cytoplasmic DNA 
viral inclusion bodies

Would like to know special stains for demonstrating intra cytoplasmic viral
inclusion bodies
tried with shorrs stain giving lot of red color
Anybody can suggest alternative method excluding immunohistochemistry!
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Re: [Histonet] Pam Plumlee wants to share new pictures with you :)

[E. Wayne Johnson 朱稳森]

Pam Plumlee thinks you're cute.


On 10/14/12 2:41, Akemi Allison wrote:

Take me off this list!  We are not a dating service!
  
Akemi Allison BS, HT(ASCP)HTL

Director
Phoenix Lab Consulting
E-Mail: akemiat3...@yahoo.com





  From: Pam Plumleepaw...@yahoo.com
To: histonet@lists.utsouthwestern.edu
Sent: Friday, October 12, 2012 5:50 PM
Subject: [Histonet] Pam Plumlee wants to share new pictures with you :)


Pam Plumlee has added you as a friend on Zoosk.

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Re: [Histonet] Traveling

[E. Wayne Johnson]

If you do that and announce your availability on Histonet, you are likely
to be criticized publicly by a few, while others send email and twitters 
to one
another privately discussing your attributes, and those who like having 
the information

that you are available probably will not speak up in your support.

But since there is no such thing as bad publicity, you will have to 
decide the +/- of it.


On 10/3/2012 5:49 AM, Demetria Ross wrote:

I'm considering becoming a traveling histotech in the future. Are there any
traveling techs or any techs that traveled in the past on the forum that
can offer some ups and downs positive and negatives of traveling Thanks.
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Re: [Histonet] microtome

[E. Wayne Johnson]

I got it open and cleaned the wheel stop mechanism and
the belt in front and it works good now and we now have a backup and
my people are smiling.

Thanks for everyone's helpful advices.

Wayne Johnson
Enruikang (Enable) Ag Tech
Beijing

On 9/22/2012 4:34 AM, Burton, Lynn wrote:

Perhaps they have a contact person since they do so much work throughout the US.

From: jsjurc...@comcast.net [mailto:jsjurc...@comcast.net]
Sent: Friday, September 21, 2012 3:16 PM
To: Jay Lundgren
Cc: Histonet; Burton, Lynn
Subject: Re: [Histonet] microtome

As I said, I'm not sure they have anybody in Beijing.

From: Jay Lundgrenjaylundg...@gmail.com
To: Lynn Burtonlynn.bur...@illinois.gov
Cc: Histonethistonet@lists.utsouthwestern.edu
Sent: Friday, September 21, 2012 2:27:59 PM
Subject: Re: [Histonet] microtome

Um.he is in Beijing.
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[Histonet] microtome

[E. Wayne Johnson]

We work in a previously underutilized histology lab.

There was a sliding horizontal Sakura microtome, a functioning Thermo 
microtome,

and a nonfunctioning Leica 5xxx (not a very fancy Leica).

The center where we work has no interest in fixing the Leica microtome.
I am supposing that it is merely dirty.  Beijing is very dusty and the 
windows to

the room leaked badly until we installed window seals.

I would like to try to take the Leica microtome apart and try to clean 
it but I cant find
the way to get it open.  There is no hatch to open like on an old AO 
Spencer.


We are told that it would cost way to much to have Leica service it.

Does anyone have any experience with maintenance or repair of a low-end 
Leica Microtome

and have any tips to share on how to open and clean it?

Wayne Johnson
Beijing.

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Re: [Histonet] microtome

[E. Wayne Johnson]

I think its a Leica 2125.

On 9/22/2012 12:32 AM, E. Wayne Johnson wrote:

We work in a previously underutilized histology lab.

There was a sliding horizontal Sakura microtome, a functioning Thermo 
microtome,

and a nonfunctioning Leica 5xxx (not a very fancy Leica).

The center where we work has no interest in fixing the Leica microtome.
I am supposing that it is merely dirty.  Beijing is very dusty and the 
windows to

the room leaked badly until we installed window seals.

I would like to try to take the Leica microtome apart and try to clean 
it but I cant find
the way to get it open.  There is no hatch to open like on an old AO 
Spencer.


We are told that it would cost way to much to have Leica service it.

Does anyone have any experience with maintenance or repair of a 
low-end Leica Microtome

and have any tips to share on how to open and clean it?

Wayne Johnson
Beijing.

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Re: [Histonet] microtome

[E. Wayne Johnson]

Tech One seems to be limited in scope to the US.

Nice hearing from you in Galesburg. I practiced pig production medicine 
in Illinois for quite a few years.

Doug Hoefling the former director and pathologist in Galesburg
is an old friend and mentor of mine, but of course he is long since retired.

Histopathology is not yet a well-developed tool for pig disease 
diagnosis in China.
We are presently one of the few diagnostic centers in China, perhaps the 
only one that routinely

does histopath on pig necropsy cases.

On 9/22/2012 4:34 AM, Burton, Lynn wrote:

Perhaps they have a contact person since they do so much work throughout the US.

From: jsjurc...@comcast.net [mailto:jsjurc...@comcast.net]
Sent: Friday, September 21, 2012 3:16 PM
To: Jay Lundgren
Cc: Histonet; Burton, Lynn
Subject: Re: [Histonet] microtome

As I said, I'm not sure they have anybody in Beijing.

From: Jay Lundgrenjaylundg...@gmail.com
To: Lynn Burtonlynn.bur...@illinois.gov
Cc: Histonethistonet@lists.utsouthwestern.edu
Sent: Friday, September 21, 2012 2:27:59 PM
Subject: Re: [Histonet] microtome

Um.he is in Beijing.
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Re: [Histonet] RE: mouse testis in Bouins

[E. Wayne Johnson]

What danger of Picric Acid are you concerned with?

Surely its not the hyped explosion hazards.

We use picric acid and as inquisitive boys we have tried very hard to 
ignite it thinking it would be fun.


We dried some down and wrapped it in aluminum foil and with appropriate 
protection outdoors beat it with a hammer.

So very disappointing.  We only made it flat.

We tried heating some.  It does burn pretty good but not really 
dramatically.
We tried purifying and recrystallizing it and it still didnt do anything 
spectacular.

Our conclusion that as fireworks, pure picric acid is pretty much a dud.

I have done some reading about picric acid and it seems that in lab 
conditions a
picric acid explosion is very unlikely maybe impossible even if the 
stuff is very dry indeed.

We do keep our picric acid wet in a safe spot for storage.

Some metal salts of picric acid are said to be much more sensitive.  We 
havent made any lead picrate to play with
since we are worried about aerosolizing the lead when it does explode or 
flash.


There are some youtube movies about how to make explosive derivatives of 
picric acid.  it seems
that picric acid is just not a very good explosive, and that small 
amounts in free open air are unlikely to explode.


I have been unable to find any reference to any lab accidents with 
picric acid.


Does anyone have any information to the contrary?




On 9/15/2012 7:55 AM, Jackie O'Connor wrote:

As a GLP tox lab, we have done away with using Bouin's altogether - there is 
literature out there (somewhere - not handy now) that indicates Modified 
Davidson's fixative provides the same testicular detail of bouins, without the 
picric acid danger.  We switched about 3-4 years ago, and our testicle experts 
are happy.  I believe most labs are getting away from Bouins.
Jackie O'


-Original Message-
From: Frances Elizabeth Barronfbar...@stanford.edu
To: histonethistonet@lists.utsouthwestern.edu
Sent: Fri, Sep 14, 2012 12:21 pm
Subject: [Histonet] RE: mouse testis in Bouins


Hi Margaret,

Our protocol for whole mouse embryos E14.5-E18.5 was to fix in Bouin's for 5-7
days at room temp (I have gone longer, but it isn't exactly recommended). Most
of the length of time, however, was to compensate for the large tissue size and
need for good penetration. I'm not sure how that converts to your particular
tissue of interest.

For long term storage, John Shelton at UT Southwestern (who did our vacuum
processing for large embryos) told me that it was preferred to put them in 1%
neutral buffered formalin and store them at room temp. We had previously been
storing them in 70% EtOH, but John said that the long exposure to EtOH leads to
excessive drying of the tissue and ultimately brittleness if used later. I'm
assuming this thought could be applied to any tissue piece, but I don't have
enough experience to really know. We have successfully gotten beautiful paraffin
sections from 3mo-1year samples that have been stored this way.

I'm hoping this will be of some help to you, and perhaps others in the list can
comment.

Best of luck,
~Francie

***

Francie Barron, Ph.D.
Postdoctoral Fellow, Joseph Wu Lab

Stanford University School of Medicine
Lorry I. Lokey Stem Cell Research Building
265 Campus Drive, Room G1105
Stanford, CA 94305-5454

Phone: (650) 724-5564 or (650) 724-9240
Fax: (650) 736-0234

***



Message: 7
Date: Fri, 14 Sep 2012 10:06:33 -0300
From: Margaret Hornemho...@upei.ca
Subject: [Histonet] mouse testis in Bouins
To:histonet@lists.utsouthwestern.edu
Message-ID:505301a902d100018...@oes-grpwise.novell.upei.ca
Content-Type: text/plain; charset=us-ascii

  Hello Everyone, I am asking this for a friend.

How long can mouse testis be kept in Bouins without distortion of cell
morphology? Days? weeks? months? years?

I noticed in the Archives that many people fix in Bouins , rinse, then
store in 70% EtOH. This is preferable I assume. Again, how long is ok?


Thanks in advance for the sharing of your accumulated wisdom,
   Margaret



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Re: [Histonet] RE: air drying special stain slides rather than

[E. Wayne Johnson]

Dishwashing machines are not at all common in China even in Beijing so
we could not find dishwasher detergent nearby.   But we took a bus and 
subway
ride toward the city center to Zhongguancun (the computer district) and 
found a Carrefour's
(Jia Le Fu 家乐福) that had one brand of powdered detergent Finish 
(Reckett Benckiser).
Finish was formerly called Electrasol.  Actually I was a bit afraid of 
Finish.  If I had known
it was the same thing as the familiar Electrasol, I would not have had 
any concerns.


Anyway, we took the Finish powder back to the lab where we had a covered 
water bath waiting at 90C.
I mixed 40 grams in with 2L of tap water and we followed the protocol 
Rene' sent with some test
tissues from some pigs we examined a few days ago (lung, liver, heart, 
spleen, kidney, gut).
We heated the detergent solution on an induction stove and poured in 
into some square glass jars in the water bath.
The procedure took the paraffin right off.  We did an HE and dried the 
slide in the 60C oven after

a water wash to clean up after Eosin.  Ver-r-ry nice result.

Jane tried the technique then by herself with 3 more slides including 
one slide with some honking big pieces of pig cerebellum.
The sections all stayed put on the slides.  Sometimes we can lose most 
of the cerebellum in processing, so
we think it is a good demonstration that section loss is not going to be 
much of a problem.
The stain was a Harris hematoxylin regressed with 1% HCl.  We blued some 
with
tap water and some with Scott's.  I sort of prefer the plain tap water 
bluing.
Our Eosin is an Eosin/Biebrich Scarlet (Acid Red 66)/Orange G that we 
make up.


We usually use a treatment in alcoholic Picric Acid to get rid of 
formalin pigment but we omitted that step today and the
slides turned out well.  Indeed these pigs were a field necropsy and the 
formalin was simple 10% unbuffered formalin in
plain local farm tap water, but there were only some traces of formalin 
pigment.  We are wondering if perhaps the detergent is

taking out some of the formalin pigment for us.

We got a few white paraffin spots on one slide but even that would not 
be an issue in reading or photographing the slide.

Jane thinks she can tweak the procedure to eliminate the paraffin spots.
Jane's opinion on the procedure?  She will be bottling up all of the 
xylenes and alcohols and storing them away first thing tomorrow

morning.

This fixes a big problem for us because the histolab is on the first 
floor along with some offices.  We do our work under
a fume hood and we are careful, but we  had an incident where students 
left containers of xylene uncapped outside the hood
overnight in hot weather vaporizing a large amount of xylene into the 
hallway.  Not cool.


We moved the tissue processor and autostainer to a remoter spot on the 
4th floor
but the water quality issues there made our autostainer a problem.  We 
can now bring our autostainer back and set it
up for special and routine stains.  The procedure with detergent from 
beginning to end
is significantly shorter than the xylene alcohols stain alcohols xylene 
procedure and we will dramatically reduce
our consumption and waste output of xylene and our consumption of 
ethanol.  Very cool.


Thanks

EW Johnson
Enruikang Ag Tech
Beijing


On 9/12/2012 3:13 AM, Rene J Buesa wrote:

EWayne:
All mounting media contains a so called solvent. Permount contains 
toluene (not as nasty as xylene) and will penetrate the section 
provided it is absolutely dehydrated (in an oven in this case).
You just have to finish the HC (or IHC) protocol and pop-in the slides 
in the oven.
Balsam of Canada (the resin) is dissolved in xylene (always) so the 
penetration is also assured.

Under separate cover I am sending you my articles.
Try this method, you will love it!
René J.

*From:* E. Wayne Johnson e...@pigsqq.org
*To:* Rene J Buesa rjbu...@yahoo.com
*Cc:* Mayer,Toysha N tnma...@mdanderson.org; 
'histonet@lists.utsouthwestern.edu' histonet@lists.utsouthwestern.edu

*Sent:* Tuesday, September 11, 2012 1:14 PM
*Subject:* Re: [Histonet] RE: air drying special stain slides rather than

I am convinced to give it a try because I also have trouble will the
loss of some stains in dehydration.
I was concerned that the slides would not clear well after oven
dehydration.  I will see how it
works for me.

I can see clearly how going from counterstain to oven will save much
hassle with xylene and alcohols as well as
not washing out some special stains.  I have tried some of the isopropyl
alcohol and acetone dehydration called for
in some of the stain procedures and it would be great if the slides
could just be popped into the oven.

What mounting medium are you using?  Does it matter?  I am a bit worried
about penetration of the mountant
into the tissue section if there is no xylene in the tissue.  Will
neutral balsam still work ok?

Rene:  if you have a link to the paper you talked about on eliminating
xylene, I am interested

Re: [Histonet] RE: air drying special stain slides rather than

[E. Wayne Johnson]

Interestingly, I showed the results to a couple of colleagues.

One response was-
The sections will come off!
The sections will come off!
All that heat!
The soap!
The sections will come off!
I don't think I even want to try That!
I'm going to stick with Xylene and Alcohols.

Another -
  Oh, this method is /Very Unusual/.

Maybe if the graduate students try to publish
a scientific paper and say that they used this method, their papers
will be *Rejected* *by the Reviewers*.

It's a published method.  I have 2 published papers on it right here

Oh, so they can cite those methods in their papers.   Ok.


*
E. Wayne Johnson
Enruikang Ag Tech
Beijing


On 9/13/2012 8:41 AM, Tony Henwood (SCHN) wrote:

Yep

Regards
Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA)
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, AUSTRALIA


-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Diana McCaig
Sent: Wednesday, 12 September 2012 3:23 AM
To: E. Wayne Johnson; Rene J Buesa
Cc: histonet@lists.utsouthwestern.edu; Mayer, Toysha N
Subject: RE: [Histonet] RE: air drying special stain slides rather than

Would this work for auto cover slipping  (tape film)if they were set in the 
xylene reservoir prior to cover slipping?

Diana

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of E. Wayne Johnson
Sent: September-11-12 1:15 PM
To: Rene J Buesa
Cc: 'histonet@lists.utsouthwestern.edu'; Mayer, Toysha N
Subject: Re: [Histonet] RE: air drying special stain slides rather than

I am convinced to give it a try because I also have trouble will the loss of 
some stains in dehydration.
I was concerned that the slides would not clear well after oven dehydration.  I 
will see how it works for me.

I can see clearly how going from counterstain to oven will save much hassle 
with xylene and alcohols as well as not washing out some special stains.  I 
have tried some of the isopropyl alcohol and acetone dehydration called for in 
some of the stain procedures and it would be great if the slides could just be 
popped into the oven.

What mounting medium are you using?  Does it matter?  I am a bit worried about 
penetration of the mountant into the tissue section if there is no xylene in 
the tissue.  Will neutral balsam still work ok?

Rene:  if you have a link to the paper you talked about on eliminating xylene, 
I am interested.  Xylene is becoming more and more of an issue and a pain for 
us.

EWJohnson
Enruikang Ag Tech
Beijing.


On 9/12/2012 12:01 AM, Rene J Buesa wrote:
   

Toysha:
Perhaps you have not oven dried stained slides before, and that explains some 
of your comments, like:
1- if the stained slides are completely dried, the miscibility you
point out is not an issues, because there is nothing to mix with;
2- if you dehydrate → clear the stained sections that will take about
15 minutes per group of up to 25 slides, or even more depending on the
protocol used in your automated stainer, but if your group of slides
in their rack are placed in an oven at 60ºC for 5 minutes it will just
that, 5 minutes reducing the usual TAT for each staining procedure;
3- any oven can accommodate more than 100 stained slides in their
racks and the TAT is shortened by oven drying, no matter how many
slides you are working with;
4- I really do not know where you can find that extreme heat can
affect the tissue sections. All tissue sections are fixed → processed
→ dried (usually at the same 60ºC before staining) → stained and an
additional step at 60ºC to dry before cover-slipping is just that, an
additional step at 60ºC
5- The so called Lean technologies do not refer to staining only,
they have to do with the whole work-flow and an additional drying step
at 60ºC cannot affect in a negative way to the work-flow
6- after staining you will oven dry the sections.
I think you should try the method instead.
René J.



From: Mayer,Toysha Ntnma...@mdanderson.org
To:
'histonet@lists.utsouthwestern.edu'histo...@lists.utsouthwestern.ed
u
Sent: Tuesday, September 11, 2012 11:41 AM
Subject: [Histonet] RE: air drying special stain slides rather than


Ooh, great question for my students next semester.
Your answer is the counterstain, some counterstains may require dehydration 
after rinsing, or some may not. Adjusting the times of the counterstain is not 
the issue as much as  the solvent of the counterstain.

Rene, while I do acknowledge that the xylene may/will cause hazards, we must 
think of the miscibility of the clearant and the dehydrant, as well as the 
amount of time involved.  The amount of time involved to blot and air dry the 
slides will affect the TAT for the specimen.  5 min may

Re: [Histonet] RE: air drying special stain slides rather than

[E. Wayne Johnson]

I am convinced to give it a try because I also have trouble will the 
loss of some stains in dehydration.
I was concerned that the slides would not clear well after oven 
dehydration.  I will see how it

works for me.

I can see clearly how going from counterstain to oven will save much 
hassle with xylene and alcohols as well as
not washing out some special stains.  I have tried some of the isopropyl 
alcohol and acetone dehydration called for
in some of the stain procedures and it would be great if the slides 
could just be popped into the oven.


What mounting medium are you using?  Does it matter?  I am a bit worried 
about penetration of the mountant
into the tissue section if there is no xylene in the tissue.  Will 
neutral balsam still work ok?


Rene:  if you have a link to the paper you talked about on eliminating 
xylene, I am interested.  Xylene is becoming

more and more of an issue and a pain for us.

EWJohnson
Enruikang Ag Tech
Beijing.


On 9/12/2012 12:01 AM, Rene J Buesa wrote:

Toysha:
Perhaps you have not oven dried stained slides before, and that explains some 
of your comments, like:
1- if the stained slides are completely dried, the miscibility you point out 
is not an issues, because there is nothing to mix with;
2- if you dehydrate → clear the stained sections that will take about 15 
minutes per group of up to 25 slides, or even more depending on the protocol 
used in your automated stainer, but if your group of slides in their rack are 
placed in an oven at 60ºC for 5 minutes it will just that, 5 minutes reducing 
the usual TAT for each staining procedure;
3- any oven can accommodate more than 100 stained slides in their racks and the 
TAT is shortened by oven drying, no matter how many slides you are working with;
4- I really do not know where you can find that extreme heat can affect the 
tissue sections. All tissue sections are fixed → processed → dried (usually at the same 
60ºC before staining) → stained and an additional step at 60ºC to dry before 
cover-slipping is just that, an additional step at 60ºC
5- The so called Lean technologies do not refer to staining only, they have 
to do with the whole work-flow and an additional drying step at 60ºC cannot affect in a 
negative way to the work-flow
6- after staining you will oven dry the sections.
I think you should try the method instead.
René J.



From: Mayer,Toysha Ntnma...@mdanderson.org
To: 'histonet@lists.utsouthwestern.edu'histonet@lists.utsouthwestern.edu
Sent: Tuesday, September 11, 2012 11:41 AM
Subject: [Histonet] RE: air drying special stain slides rather than


Ooh, great question for my students next semester. 
Your answer is the counterstain, some counterstains may require dehydration after rinsing, or some may not. Adjusting the times of the counterstain is not the issue as much as  the solvent of the counterstain.
  
Rene, while I do acknowledge that the xylene may/will cause hazards, we must think of the miscibility of the clearant and the dehydrant, as well as the amount of time involved.  The amount of time involved to blot and air dry the slides will affect the TAT for the specimen.  5 min may be ok if you have a small amount of slides, but with a larger number of slides, it will be considerably more than 5.  Also Lean methodologies would not apply in that case. With automation, the extreme heat involved with a stain dryer may affect the tissue on the slide.


There are some stains that can be blotted, cleared and coverslipped, but using 
the alcohol to remove excess water and counter stain is better in my opinion.


Toysha N. Mayer, MBA, HT (ASCP)
Instructor, Education Coordinator
Program in Histotechnology
School of Health Professions
MD Anderson Cancer Center
(713) 563-3481
tnma...@mdanderson.org




Message: 16
Date: Tue, 11 Sep 2012 10:32:08 -0400
From: Diana McCaigdmcc...@ckha.on.ca
Subject: [Histonet] air drying special stain slides rather than
 dehydrateand clear
To:histonet@lists.utsouthwestern.edu
Message-ID:
 dcfd9e6a390e294aaf3a2561cd32e5c417a90...@ckhamail1.ckha.on.ca
Content-Type: text/plain;charset=us-ascii

I was hoping to get information on why special stains are dehydrated, cleared 
and mounted vs allowing them to be blotted dry, air dried then coverslip.



Every procedure I have ever encountered always indicates to dehydrate and clear 
but I have heard where some labs are blotting the slides , allowing to air dry 
(probably not set standard time) and dipped in xylene prior to cover slipping.  
Reason given is that the counterstain gets washed out.  Wouldn't adjusting the 
times be a better resolution.



I understand residual water could be present and cause long term issues on 
storage but wanted some other opinions on this process.



Diana



--

Message: 17
Date: Tue, 11 Sep 2012 07:52:05 -0700 (PDT)
From: Rene J Buesarjbu...@yahoo.com
Subject: Re: [Histonet] air drying special stain slides rather than
 

Re: [Histonet] annoying crystals on sections

[E. Wayne Johnson]

we are using a Sakura DRS2000 and we are 3 x 3 minutes in xylene and we have
been keeping it fresh.  The staining is good now but we still see the 
crystals.

If it were paraffin we should see unstained spots on the slide I think.

I have gone to an aqueous 1% HCl today after hematoxylin for regression 
and that seems to be cleaning
them up on most of the slides.  I cleaned out some of the plumbing and 
cleaned some calcium out of the pipes.


We are using Harris hematoxylin that we purchase.  We have a  tried 
different counterstains but it seems to

make no difference.

We are using a Sakura tissue processor for overnight processing of 
cassettes.  the embedding is going good
and we get nice flat thin sections.  We are fixing tissues with neutral 
phosphate buffered formalin but
still see some formalin pigment.  We are cleaning that up with picric 
acid in etoh.  We find we still need that
and the formalin pigment is brown to dark brown.  These problem crystals 
are round irregular to rhomboidal
some times sort of large and flat about the size of a cell and they are 
clear.  I thought they were formalin pigment
at first and fiddled with the Picric Acid, and tried Ammonia in Alcohol 
to get rid of formalin pigment and finally

decided that it was not formalin pigment.

I thought it might be something from Scott's tap water (Mg++) so i 
dropped that and tried bluing with NH4+
and it didnt help any.  I tried blueing just with tap water.  Nice 
result but still the crystals.


The aqueous HCl seems to be working and is not harming the nuclei so I 
may have a sort of solution

and am calling it calcium crystals in the water until I know better.

I may look for some sort of filter to put in the water line.

E Wayne Johnson DVM
Enruikang Ag Tech
MOA Feed Industry Centre
China Agriculture University
Beijing

On 8/21/2012 7:51 PM, Debra Siena wrote:

could it be paraffin?
Debbie Siena HT(ASCP)QIHC
Technical Manager | StatLab Medical Products
407 Interchange St. | McKinney, TX 75071
Direct: 972-436-1010  x229 | Fax: 972-436-1369
dsi...@statlab.com | www.statlab.com


- Original Message -
From: e...@pigsqq.org [mailto:e...@pigsqq.org]
Sent: Tuesday, August 21, 2012 05:46 AM
To: Debra Siena
Subject: Re: [Histonet] annoying crystals on sections

We just now ran the statlab version protocol from StatLab's website, using an 
alcoholic eosin.  No doubt that gives a stronger brighter red stain.  However 
we still see those crystals!

We are suspecting a water problem.  I have been dismantling some of the 
plumbing and getting some Ca crystals out of the pipes.

We are manually coverslipping.

   

  ---Original Message---
  From: Debra Sienadsi...@statlab.com
  To: 'e...@pigsqq.org'e...@pigsqq.org
  Subject: Re: [Histonet] annoying crystals on sections
  Sent: Aug 21 '12 07:46

  Is your eosin alcoholic or aqueous?  What is your staining protocol and what reagents are you using?  This information would be most helpful. Also are the sections paraffin embedded and routinely processed in tissue processor?  
  Debbie Siena HT(ASCP)QIHC

  Technical Manager | StatLab Medical Products
  407 Interchange St. | McKinney, TX 75071
  Direct: 972-436-1010  x229 | Fax: 972-436-1369
  dsi...@statlab.com | www.statlab.com


  - Original Message -
  From: E. Wayne Johnson [mailto:e...@pigsqq.org]
  Sent: Monday, August 20, 2012 06:20 PM
  To: histonet@lists.utsouthwestern.eduhistonet@lists.utsouthwestern.edu
  Subject: [Histonet] annoying crystals on sections

  We are having problems with crystals precipitated on our slides which
  are HE stains on tissues from pigs.
  Tissues are fixed in buffered formalin.  We had trouble months ago with
  formalin pigment and we had resolved
  that by using ammonia in EtOH or picric acid in EtOH.  Sometimes we
  receive fixed samples from the field that are
  not buffered but presently all of our tissues are fixed in neutral
  phosphate buffered formalin.

  We moved the Sakura autostainer to a different location under a fume
  hood on a
  different floor of the building to get the solvent odor out of our work
  area.
  Immediately we began to see a tremendous degradation in slide quality
  due to what we initially thought was formalin pigment.
  We have changed all of the solutions and all of the stains.  We find
  that if we use Milli-q water instead of tap water for
  rinsing (done by hand in that case) we dont see the crystals, but the
  eosin staining quality is not acceptable after rinsing in the acidic
  (ph ~5) Milli-Q water.

  Our tap water is neutral to slightly alkaline and is very hard with calcium.

  We do all sorts of tissues for diagnosis of pig diseases.  Sometimes the
  slides are quite acceptable but sometimes particularly
  when looking at small intestine, the crystals are very annoying.  The
  crystals occur randomly on the slide except that there is a
  tendency for them to be centered on nuclei particularly in intestinal
  epithelium.  The crystals

[Histonet] Re: annoying crystals on sections

[E. Wayne Johnson]

Thanks all for many useful and helpful suggestions
and interesting anecdotes.


E. Wayne Johnson, DVM
Enruikang AgTech
MOA Feed Industry Centre
China Agricultural University
Beijing





On 8/22/2012 5:27 AM, David A. Wright wrote:

Hi Wayne  Histonet

My guess is that Wayne's crystals are Calcium phosphate - the calcium from the 
hard tap water, as described, and phosphate from phosphate buffer (sodium or 
potassium) in the solution immediately preceding the tapwater rinse. The 
variation in crystal deposition would then be in the degree each tissue/ 
organelle tends to carry over the phosphate into the tapwater wash. Just mix 
drops of the solutions together on a slide and see if crystals form.

A brief deionized rinse followed by tapwater should first remove the phosphate 
(  crystals) and then allow the desired blueing. Alternatively, substitute 
TRIS or similar as the buffer in the preceding step and go directly to tapwater.

If you have valuable sections with crystals on them, you should be able to 
chelate away the deposits in an EDTA solution, then restain as needed.
  all the best!
-David
==
David A. Wright, Ph.D.
University of Chicago Section of Neurosurgery

 Original message 
   

Date: Tue, 21 Aug 2012 09:44:26 -0500 (CDT)
Subject: Histonet Digest, Vol 105, Issue 25 Message: 12
Date: Tue, 21 Aug 2012 07:20:06 +0800
From: E. Wayne Johnsone...@pigsqq.org
Subject: [Histonet] annoying crystals on sections

We are having problems with crystals precipitated on our slides which are HE 
stains on tissues from pigs. Tissues are fixed in buffered formalin.  We had 
trouble months ago with formalin pigment and we had resolved that by using ammonia 
in EtOH or picric acid in EtOH.  Sometimes we receive fixed samples from the field 
that are not buffered but presently all of our tissues are fixed in neutral 
phosphate buffered formalin.

We moved the Sakura autostainer to a different location under a fume hood on a 
different floor of the building to get the solvent odor out of our work area.
Immediately we began to see a tremendous degradation in slide quality due to 
what we initially thought was formalin pigment.
We have changed all of the solutions and all of the stains.  We find that if we 
use Milli-q water instead of tap water for
rinsing (done by hand in that case) we don't see the crystals, but the eosin 
staining quality is not acceptable after rinsing in the acidic (ph ~5) Milli-Q 
water.

Our tap water is neutral to slightly alkaline and is very hard with calcium.

We do all sorts of tissues for diagnosis of pig diseases.  Sometimes the slides 
are quite acceptable but sometimes particularly when looking at small 
intestine, the crystals are very annoying.  The crystals occur randomly on the 
slide except that there is a tendency for them to be centered on nuclei 
particularly in intestinal epithelium.  The crystals are birefringent in 
polarized light but seem to be generally clear not dark like the formalin 
pigment we had seen before.  Neither ammonia nor picric acid remove these, and 
now if we use alcoholic ammonia to treat the slides, the slides come out too 
blue.  Our slides are cut at 4 to 5 microns.

Brain has the least problem, small intestine seems worst.

We have gone back and cut some blocks that previously stained beautifully with 
no pigment or precipitate problems
and those slides also now have the same problem, either crystals if washed with 
tap water, or poor eosin staining if rinsed with MilliQ water.

Our next step is to examine the slides microscopically at every step and try to 
find at which step the problem is occurring.

Any thoughts or similar experiences?

E. Wayne Johnson, DVM
Enruikang AgTech
MOA Feed Industry Centre
Beijing
--
 


   


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Histonet mailing list
Histonet@lists.utsouthwestern.edu
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[Histonet] annoying crystals on sections

[E. Wayne Johnson]

We are having problems with crystals precipitated on our slides which 
are HE stains on tissues from pigs.
Tissues are fixed in buffered formalin.  We had trouble months ago with 
formalin pigment and we had resolved
that by using ammonia in EtOH or picric acid in EtOH.  Sometimes we 
receive fixed samples from the field that are
not buffered but presently all of our tissues are fixed in neutral 
phosphate buffered formalin.


We moved the Sakura autostainer to a different location under a fume 
hood on a
different floor of the building to get the solvent odor out of our work 
area.
Immediately we began to see a tremendous degradation in slide quality 
due to what we initially thought was formalin pigment.
We have changed all of the solutions and all of the stains.  We find 
that if we use Milli-q water instead of tap water for
rinsing (done by hand in that case) we dont see the crystals, but the 
eosin staining quality is not acceptable after rinsing in the acidic

(ph ~5) Milli-Q water.

Our tap water is neutral to slightly alkaline and is very hard with calcium.

We do all sorts of tissues for diagnosis of pig diseases.  Sometimes the 
slides are quite acceptable but sometimes particularly
when looking at small intestine, the crystals are very annoying.  The 
crystals occur randomly on the slide except that there is a
tendency for them to be centered on nuclei particularly in intestinal 
epithelium.  The crystals are birefringent in polarized light but
seem to be generally clear not dark like the formalin pigment we had 
seen before.  Neither ammonia nor picric acid remove these,
and now if we use alcoholic ammonia to treat the slides, the slides come 
out too blue.  Our slides are cut at 4 to 5 microns.


Brain has the least problem, small intestine seems worst.

We have gone back and cut some blocks that previously stained 
beautifully with no pigment or precipitate problems
and those slides also now have the same problem, either crystals if 
washed with tap water, or poor eosin staining if rinsed with MilliQ water.


Our next step is to examine the slides microscopically at every step and 
try to find at which step the problem is occurring.


Any thoughts or similar experiences?

E. Wayne Johnson, DVM
Enruikang AgTech
MOA Feed Industry Centre
Beijing

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