Re: [Histonet] IHC-H HELP...

2019-01-03 Thread Tony Henwood (SCHN) via Histonet 
Yep,

Definitely an issue.

You can easily stain the IPX slides with H, though the discernibility of the 
cells and tissue structure with the H will depend on the degree of DAB 
product laid down (eg I would expect it to be difficult with a Vimentin IPX 
compared to a CD15).

(Grosset, A. A., Loayza-Vega, K., Adam-Granger, É., Birlea, M., Gilks, B., 
Nguyen, B., ... & Trudel, D. (2017). Hematoxylin and Eosin Counterstaining 
Protocol for Immunohistochemistry Interpretation and Diagnosis. Applied 
immunohistochemistry & molecular morphology: AIMM.)

As for doing another IPX on the existing IPX stained section (with DAB as the 
chromogen), you will have to use a different label (eg alkaline phosphatase). 
The result will depend on the cell compartment the two antigens exist. If the 
DAB is nuclear, then a cytoplasm or cytoplasmic membrane localisation with the 
Alk Phosphatase will work. If both antigens are cytoplasmic, then you will not 
see co-localisation in the same cell since the DAB will prevent any antibody 
binding in the same compartment.

Assuming the above is good, then since the antigen retrieval tends to reverse 
over time, I would include a short retrieval before the second antibody.

Now after all that, I hope the section stays on the slide!


Regards 

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) 
Principal Scientist, the Children's Hospital at Westmead
Adjunct Fellow, School of Medicine, University of Western Sydney 
Tel: 612 9845 3306 
Fax: 612 9845 3318 
Pathology Department
the children's hospital at westmead
Cnr Hawkesbury Road and Hainsworth Street, Westmead
Locked Bag 4001, Westmead NSW 2145, AUSTRALIA 


-Original Message-
From: Curt via Histonet [mailto:histonet@lists.utsouthwestern.edu] 
Sent: Friday, 4 January 2019 4:05 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] IHC-H HELP...

Need help before I turn a mistake into an irreparable mistake...

We have some unstained slides that were supposed to get stained with H but my 
guy stained them with IHC. It's complicated, we received slides and a block, 
the block was for IHC, the unstained slides were for H, he inverted the 
process) The point is, now the unstained slides are stained with IHC... I know 
we cannot destain the IHC but we can simply run and H over them... the real 
question I have is subsequent to the H this pathologist generally likes to 
see the H then order IHC on them based on what he sees (we only have these 
few unstained slides, don't have blocks to recut)...

So the question is... if we've already run IHC, then followed that with and 
H, can we return to run IHC on the slides again? would you want to skip any 
pre-treatment, antigen retrieval

I don't see this working too well myself, if they're already stained with DAB, 
that would be present on the second stain...

Thoughts?

Thanks for your help.

Curt

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[Histonet] Xylene substitute

2019-01-03 Thread Logan, Shannon via Histonet 
Hi Heather,

We have been using Formula 83 by CBG Biotech for over 7 years now with no 
problems, we even
recycle it.



Shannon H. Logan  B.S., HTL (ASCP)
Pathology Department

Bellin Health Memorial Hospital
744 South Webster Avenue
Green Bay, WI 54305-3400
920-433-3653  X3727

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Re: [Histonet] Vendor with all items needed/ Histonet Digest, Vol 182, Issue 3

2019-01-03 Thread John O'Brien via Histonet 
Hi Heather,
 Please contact IMEB INC for pricing and service on all your pathology
supplies, chemical needs and Pathology instruments.
We will provide the service you require to keep cost down,
Contact tra...@imebinc.com
Or   sa...@imebinc.com
Web site  www.imebinc.com
Regards,
IMEB INC

-Original Message-
From: histonet-requ...@lists.utsouthwestern.edu
[mailto:histonet-requ...@lists.utsouthwestern.edu] 
Sent: Thursday, January 03, 2019 10:00 AM
To: histonet@lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 182, Issue 3

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Today's Topics:

   1. IHC Negative reagent controls (Cayman Fleck)
   2. Re: IHC Negative reagent controls (Tony Henwood (SCHN))
   3. AM to statlab and recommendations (Heather Marlatt)
   4. Re: IHC Negative reagent controls (John Garratt)
   5. IHC-H HELP... (Curt)


--

Message: 1
Date: Thu, 3 Jan 2019 02:48:10 +
From: Cayman Fleck 
To: "histonet@lists.utsouthwestern.edu"

Subject: [Histonet] IHC Negative reagent controls
Message-ID:



Content-Type: text/plain; charset="iso-8859-1"

A question that came up regarding negative reagent controls for
IHC...currently using Ventana i-View.  Our regular negative control goes
through the standard antigen retrieval steps, like 99% of our antibodies.
However there are a small number of antibodies that require enzyme as well
(Protease 1).  I've seen a number of suggestions regarding this for the
negative reagent control...some say use an additional negative control
protocol that includes the protease, some say to use a single negative
control protocol and just include the harshest cell conditioning that any of
your protocols use (so basically use the cell conditioning + protease
negative control for all antibodies)...i-View is not polymer-based so we
need to continue using negative controls.  Any thoughts or advice?

Frank

Sent from Outlook


--

Message: 2
Date: Thu, 3 Jan 2019 04:06:57 +
From: "Tony Henwood (SCHN)" 
To: Cayman Fleck 
Cc: "histonet@lists.utsouthwestern.edu"

Subject: Re: [Histonet] IHC Negative reagent controls
Message-ID:
<0e5c3ac509b7424e942160149bb36...@svdcmbx-mex024.nswhealth.net>
Content-Type: text/plain; charset="us-ascii"

Hi Cayman,

Unfortunately, applying HIER to a negative control for an antibody that
requires enzyme retrieval (or no retrieval at all) is not appropriate.
The pre-treatment processes are different and could unmask different
epitopes.
If you are using a negative control then the whole procedure needs to be
same with the exception or replacing the localisation antibody with an
Isotypic antibody solution. (Isotype controls are primary antibodies that
lack specificity to the target, but match the class and type of the primary
antibody used in the application.)

For example, applying citrate or EDTA HIER to sections prior to using the
CD99 antibody (clone 12E7) can reveal perinuclear (golgi-like) staining of
some tumours (eg some colonic carcinomas) but this is not seen if enzyme
retrieval is used.

Hope this is useful

Regards 

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA)
Principal Scientist, the Children's Hospital at Westmead Adjunct Fellow,
School of Medicine, University of Western Sydney
Tel: 612 9845 3306
Fax: 612 9845 3318
Pathology Department
the children's hospital at westmead
Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001,
Westmead NSW 2145, AUSTRALIA 




-Original Message-
From: Cayman Fleck via Histonet [mailto:histonet@lists.utsouthwestern.edu]
Sent: Thursday, 3 January 2019 1:48 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] IHC Negative reagent controls

A question that came up regarding negative reagent controls for
IHC...currently using Ventana i-View.  Our regular negative control goes
through the standard antigen retrieval steps, like 99% of our antibodies.
However there are a small number of antibodies that require enzyme as well
(Protease 1).  I've seen a number of suggestions regarding this for the
negative reagent control...some say use an additional negative control
protocol that includes the protease, some say to use a single negative
control protocol and just include the harshest cell conditioning that any of
your protocols use (so basically use the cell conditioning + protease
negative control for all

[Histonet] IHC-H HELP...

2019-01-03 Thread Curt via Histonet 
Need help before I turn a mistake into an irreparable mistake...

We have some unstained slides that were supposed to get stained with H but my 
guy stained them with IHC. It's complicated, we received slides and a block, 
the block was for IHC, the unstained slides were for H, he inverted the 
process)
The point is, now the unstained slides are stained with IHC... I know we cannot 
destain the IHC but we can simply run and H over them... the real question I 
have is subsequent to the H this pathologist generally likes to see the 
H then order IHC on them based on what he sees (we only have these few 
unstained slides, don't have blocks to recut)...

So the question is... if we've already run IHC, then followed that with and 
H, can we return to run IHC on the slides again? would you want to skip any 
pre-treatment, antigen retrieval

I don't see this working too well myself, if they're already stained with DAB, 
that would be present on the second stain...

Thoughts?

Thanks for your help.

Curt

CONFIDENTIALITY NOTE: The information transmitted, including attachments, is 
intended only for the person(s) or entity to which it is addressed and may 
contain confidential and/or privileged material. Any review, retransmission, 
dissemination or other use of, or taking of any action in reliance upon this 
information by persons or entities other than the intended recipient is 
prohibited. If you received this in error, please contact the sender and 
destroy any copies of this information.
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Re: [Histonet] IHC Negative reagent controls

2019-01-03 Thread John Garratt via Histonet 
The article referenced below may be of interest.

Standardization of Negative Controls in Diagnostic Immunohistochemistry: 
Recommendations From the International Ad Hoc Expert Panel
https://www.researchgate.net/publication/261516067_Standardization_of_Negative_Controls_in_Diagnostic_Immunohistochemistry_Recommendations_From_the_International_Ad_Hoc_Expert_Panel


John

www.ciqc.ca

‐‐‐ Original Message ‐‐‐
On Wednesday, January 2, 2019 6:48 PM, Cayman Fleck via Histonet 
 wrote:

> A question that came up regarding negative reagent controls for 
> IHC...currently using Ventana i-View. Our regular negative control goes 
> through the standard antigen retrieval steps, like 99% of our antibodies. 
> However there are a small number of antibodies that require enzyme as well 
> (Protease 1). I've seen a number of suggestions regarding this for the 
> negative reagent control...some say use an additional negative control 
> protocol that includes the protease, some say to use a single negative 
> control protocol and just include the harshest cell conditioning that any of 
> your protocols use (so basically use the cell conditioning + protease 
> negative control for all antibodies)...i-View is not polymer-based so we need 
> to continue using negative controls. Any thoughts or advice?
>
> Frank
>
> Sent from Outlookhttp://aka.ms/weboutlook
>
> Histonet mailing list
> Histonet@lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet



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[Histonet] AM to statlab and recommendations

2019-01-03 Thread Heather Marlatt via Histonet 
Did anyone else use American Mastertech and is now super frustrated with
Stat Lab???

I’ve used AM for years and loved them, great prices and fast shipping, now
cost is through the roof and they’re shipping all my stuff separately
costing a fortune!

Looking for recommendations on xylene alternatives and safe mounting
medium. I’ve been using transcend and cover safe two American Mastertech
signature products but I’m running for my life.

I’ll be contacting a few of the other vendors I use as well but I’d like
some recommendations on xylene alternatives we’ve been xylene free for 4
years and I’m not going back.

Thank you
Heather
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