Re: [Histonet] Validating a new manual special stain

[Beth Cox via Histonet]

The new CAP Checklist (published 6/4/2020) no longer requires 
validations for "/LDTs that employ the following methods: manual 
microscopy eg. histopathologic and cytopathologic microscopic 
interpretation/"  (see COM.40350, note 8).  So, CAP does not require it 
and won't have guidance for that.  HOWEVER, CLIA still requires 
validations!   The common way to do that is to run a minimum of 5, and 
preferably 10, different samples by the new method and grade them 
according to expected results.  A few of the samples should be 
'negative' samples.  Using various control tissue for your validations 
is perfectly acceptable.  Write that up, have the medical director 
review the slides and sign and date the bottom of the form: "This 
procedure has been validated and is approved for patient use."


Beth Cox, HTL/SCT(ASCP)QIHC
Pathology Solutions
Prescott, MI


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Message: 1
Date: Fri, 6 Nov 2020 18:35:25 +
From: "Pairan, Kelly" 
To: "'histonet@lists.utsouthwestern.edu'"

Subject: [Histonet] Validating a new manual special stain
Message-ID:
<49e4f7123d03401385e3727519640...@l1perdwmbx04.childrensroot.net>
Content-Type: text/plain; charset="us-ascii"

Good Afternoon,
We are currently updating one of our manual special stain procedures.  How have 
you documented the validating of a new special stain method at your 
institution?  We have a IHC validation protocol that follows the CAP guidelines 
but the CAP checklist does not mention special stain validations.

Thanks for your input,
Kelly


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Re: [Histonet] Cytology (Special stains & IHC)

[Beth Cox via Histonet]

Hi Karen,

The best resource for Cyto questions would probably be a recent edition 
of the Carson Histotechnology book.  The Fourth edition has some info on 
special stains for Cyto, but the new Fifth edition is expanded on that 
topic and has more info on IHCs for Cyto (Chapter 17).


For _special stains_, you would definitely want them fixed in 95% 
alcohol.  The only exception to that would be fat stains which would of 
course require air-dried slides.  It would likely be advantageous to use 
charged slides, but that is not required. Remember when special staining 
Cyto slides fixed in 95% alcohol, that the deparaffinization steps at 
the beginning of the procedure must be skipped.  You are normally okay 
with using regular FFPE controls with your Cytospins or ThinPrep special 
stains.


Doing _IHCs_ on Cytology preps (Cytospin or ThinPrep) gets a little 
trickier. _IF_ you are going to do them, you would want them to be 95% 
alcohol fixed and on charged slides.  However, you would be required to 
do a validation for Cytology for any/all antibodies you were going to 
run.  (Your CAP or CLIA inspector will eat you alive if you don't!).  
Alcohol fixed specimens will require a new optimization since the 
pretreatments will be different than those for FFPE.   Many antibodies, 
particularly nuclear ones, may be difficult or impossible to stain on 
the intact cells in a Cyto prep.


Also, you would not be able to use FFPE controls for Cyto IHC since they 
are 'fixed and processed differently" than the Cyto slide and therefor 
don't properly control the process.  AND, alcohol fixed smears begin to 
lose their antigenicity in about 24 hours, so they must be stained the 
same day the slides are made, so having a control bank of alcohol fixed 
slides isn't feasible.


Doing IHC on Cyto smears using FFPE protocols and FFPE controls is very 
likely to give you false negative results.  Unless you are doing large 
volume Cyto, it is very strongly recommended that you do your IHCs for 
Cyto on the FFPE cell block.


Beth Cox, HTL/SCT(ASCP)QIHC

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Message: 5
Date: Wed, 5 Aug 2020 15:34:28 +
From: "Heckford, Karen - SMMC-SF"
To:"histonet@lists.utsouthwestern.edu"

Subject: [Histonet] Cytology
Message-ID:<903be99abb344593a73079761d762...@phx-exch-013.chw.edu>
Content-Type: text/plain; charset="us-ascii"

Good Morning,
I rarely do Special stains or IHC's on Cytology cytospins or thin prep type 
slides.Should these slides be charged, air dried or fixed in 95% alcohol 
before doing Special stains or IHC's on them?

Is there a good book or some sort of publication on cytology procedures 
regarding the above mentioned.   I have tried and look it up and it seems all 
over the place on what to do.

Any help would be greatly appreciated,

Karen Heckford HT ASCP CE
Lead Histology Technician
St. Mary's Medical Center
450 Stanyan St.
San Francisco, Ca. 94117
415-668-1000 ext. 6167
karen.heckf...@dignityhealth.org

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Re: [Histonet] plural fluid prep

[Beth Cox via Histonet]

Hi Michelle,

When I was in my Cytology training, we did an experiment to answer just 
that question.  We started with a 2 liter jug of plural fluid with 
nothing added, and we stored in in the fridge.  At first we tested it 
every day to see if there was any degeneration, then we got tired of 
that and tested twice a week, then once a week. After 4 months it was 
still just fine, and we threw it away because we were tired of testing 
it (and it was the end of the semester!).   It seems that the cells are 
happy in their natural habitat and refrigeration slowed/prevented any 
bacterial growth.


So  my advice is:  don't add anything to it, keep it in the fridge, and 
it will last many months!!


Beth Cox, SCT/HTL(ASCP)QIHC

From: Michelle Jamison
To:"histonet@lists.utsouthwestern.edu"
   
Subject: [Histonet] plural fluid prep
Message-ID:
Content-Type: text/plain; charset=utf-8; format=flowed

Hi All,
I work in a research lab where we are using plural fluid to proof a
method. Recently we have received some, and are unsure how long it will
be ok in the fridge, or if we should add some preservation fluid to it.
Can anyone advise please.


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Re: [Histonet] Histonet,

[Beth Cox via Histonet]

Lisa,

I have sent you a booklet on detailed Cell Block procedures as an 
attachment in a separate email --  if anyone else is interested in a 
copy, please email me directly.


Thanks

Beth Cox, SCT/HTL(ASCP)QIHC


--

Message: 1
Date: Wed, 25 Apr 2018 17:11:43 +
From: "White, Lisa M."
To:"histonet@lists.utsouthwestern.edu"

Subject: [Histonet] Cell Block
Message-ID:



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

Does anyone have the recipe for making cell blocks with Thrombin and Plasma?
We made them at my first Histo job, oh say over 20 years ago but I do not 
remember the ratio.

Thanks bunches,
Lisa


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Re: [Histonet] Pap stain without xylene

[Beth Cox via Histonet]

Mike,

As an alternative for your discussion:

I work on medical mission trips where I run Paps in a field setting.  For the 
past 4 years, we have not had access to xylene, and we have done the following:

1. Remove slides from the last alcohol after staining
2. Allow to air dry completely (generally only takes a few minutes)
3. Use standard mounting media to coverslip them dry.

This works beautifully.  Microscopically they look the same as xylene cleared 
slides.  The only caveat is that they must dry completely.

--
*BETH COX, HTL/SCT(ASCP)QIHC*
AP Consultant | Pathology Solutions, Inc | (810) 240-2190 | bethc...@gmail.com


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Message: 6
Date: Tue, 24 May 2016 17:08:51 -0500
From: Mike Toole
To:"histonet@lists.utsouthwestern.edu"

Subject: [Histonet] Pap stain without xylene
Message-ID: <31530E35E0BAB044B3B56B7FE5CF4EB33E1B2A3240@mail>
Content-Type: text/plain; charset="iso-8859-1"

Ren? and other Histonetters,


After reading the paper:



Buesa RJ, Peshkov, MV. Histology without xylene. Ann Diagn Pathol. 2009 
Aug;13(4):246-56. Epub 2009 Feb 5.



It appears that xylene in the final clearing steps is replaced with isopropanol 
and mineral oil as follows:

?5:1 isopropanol to mineral oil 50?C

?2:1 isopropanol to mineral oil 50?C

?Undiluted mineral oil 50?C

?Drying oven 5 minutes   60?C

?Coverslip



Do you feel these clearing steps be applied to the pap stain in order to 
eliminate xylene?  If so, can it be done at room temperature?

Thanks,
Mike

Mike Toole, BS, CT(ASCP)CM


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[Histonet] PAP staining quality

[Beth Cox via Histonet]

Hi Charles,

A couple things to check on:

1.  The first concern I would have is your EA stain.  Poor EA staining 
will give too much orange staining and pale other counterstains (making 
them look aged).What brand are you using? Have you changed brands? 
Is your EA close to the expiration date? Is the bulk stored with light 
exposure?  I think fixing your EA will fix all the other problems.


2.  The other question I have regards your alcohol.  Have you changed 
types/brands?  Pap staining is very delicate and the different alcohols 
used can make a big difference.


Beth Cox, HTL/SCT(ASCP)QIHC

--
Message: 3 Date: Thu, 4 Feb 2016 09:50:18 -0500
From: Charles Riley 
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] PAP stain quality
Message-ID: 


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

Not sure if anyone out the would know the answer to this. We are having 
an issue with our PAP stained slides appearing too orange and look aged. 
If you have any idea for causes I appreciate any help


-- Charles Riley HT(ASCP)CM
Histopathology Coordinator/ Mohs
 Doctors Pathology Services, Dover DE

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Re: [Histonet] Pathology labs using Premier for staffing

[Beth Cox via Histonet]

The way Premier can break down the productivity/staffing info for your 
lab is dependent on how your organization separates the lab into cost 
centers -- if your lab is all one cost center for clinical and anatomic 
labs together, that's the way it gets reported to Premier, and that's 
how the statistics will come back to you.  These statistics are based on 
billable tests and worked hours for your cost center.  If your 
organization has the anatomic lab as a different cost center from the 
clinical lab, the info will get reported to Premier separately and 
Premier can give you info back comparing you to other AP only 
databases.   There are no labs I am aware of which have Histo and Cyto 
in separate cost centers, so Premier would likely be unable to give you 
comparison data that way.


Beth Cox, HTL/SCT(ASCP)QIHC
Pathology Solutions, Inc

Message: 10
Date: Thu, 15 Oct 2015 11:18:39 -0400
From: Amy Self
To:"histonet@lists.utsouthwestern.edu"

Subject: [Histonet] Pathology labs using Premier for staffing
Message-ID:

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


Dear Histonetters,

Are there any labs that utilize Premier for staffing productivity?   For those 
that do use Premier is the histology section separated from your other lab 
sections or included with the clinical lab?  And also is cytology considered 
separate or included with histology?  Do you report volume and staffing?   Or, 
only report one or the other?

I am trying to find a comparison group for staffing productivity. Our histology 
and cytology sections are included as part of the entire lab for both volume 
and staffing.


Thanks in advance for your help,
Amy Self
Histology Lab Senior Tech
Lab
Tidelands Georgetown Memorial Hospital
606 Black River Road
Georgetown, SC 29440
843-520-8711
as...@tidelandshealth.org


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