[Histonet] RE: MOHS IPs

Wed Oct 2 09:47:26 CDT 2013

Thanks for the reply,

We are pre-fixing in a formalin substitute, but we'll give the acetone a try.  I have diluted the protease, but have not gotten any good results.  We'll keep trying with that as well.

We've tried multiple incubation times for the cytokeratin... From 5 minutes all the way up to 30 minutes... With no luck.

We are using an enhanced polymer DAB.

Thanks for the suggestions!  I greatly appreciate it!  :)

Karen

Karen L. Bauer HTL/HT (ASCP) | Histology Supervisor | Pathology | MOHS Lab Supervisor | Dermatology | Phone: 715-838-3205 | bauer.karen <@t> mayo.edu | Mayo Clinic Health System | 1221 Whipple Street | Eau Claire, WI 54702 | mayoclinichealthsystem.org

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of pruegg <@t> ihctech.net
Sent: Wednesday, October 02, 2013 9:36 AM
To: Barbara Tibbs; Bauer, Karen L.; 'histonet <@t> lists.utsouthwestern.edu'
Subject: RE: [Histonet] RE: MOHS IPs

   I  agree with Barbara try fixing in cold acetone/ethanol mix even =
   just  for  a  couple  of minutes then go directly to buffer do not dry
   after=x,  I  would dilute the protease 1:4 with buffer (PBS or TBS
   what  ever  u  u=) and do it for a short time.  Enzyme digestion on
   frozen  sections  i=ricky  but  fixing a little and diluting the
   enzyme  should  help  prevent  ov=  digestion.   How  long  are  u
   incubating  the  primary?   Are  u usin=n enhanced labeled polymer
   detection?  HRP/Dab or AEC or Alk.P/fast=d?

-------- Original Message --------

Subject: [Histonet] RE: MOHS =s

From: Barbara Tibbs <[1]barbara.tibbs <@t> accuratediagnosticlabs.com>

D=e: Wed, October 02, 2013 5:54 am

To: "Bauer, Karen L." <[2]Bauer.Karen <@t> mayo.edu>,

"'histonet <@t> lists.utsouthwest=n.edu'"

<[3]histonet <@t> lists.utsouthwestern.edu>

Hello Karen,
   =A

Back  in the old days of doing IHC manually on mostly fresh, froze   n  tissue  I would immediately place the slide with the frozen section
   into  a=plin  jar  of  acetone  to  fix it. Keep the coplin jar of
   acetone  in  the  cr=stat.  Leave  it  in  the acetone for only 10
   minutes.  Try diluting the Pr=ease 2 - 1:1 to make it gentler so it
   doesn't  "eat"  the  tissue.  This  te=nique worked well when I was
   doing ER/PR on frozen breast tumors. Not su= it will work with skin
   but it's worth a try.

Barbara S. Tib=

Histology Supervisor

Accurate Diagnostic Labs

South Pl=nfield, NJ

[4]barbara.tibbs <@t> accuratediagnosticlabs.com

732-839-3374

C=l: 610-809-6508

_____________________________________=_

From:                  [5]histonet-bounces <@t> lists.utsouthwestern.edu
   <[6]histonet-bounces <@t> lists.utsouthwestern=du>  on behalf of Bauer,
   Karen L. <[7]Bauer.Karen <@t> mayo.edu>

Sent: Tuesday, October 01, 20 5:22 PM

To: '[8]=stonet <@t> lists.utsouthwestern.edu'

Subject: [Histonet] MOHS IPs

Hi all,

We  are  in the process of validating some a=ibodies in our MOHS
   lab.  The Melan A (Mart 1) antibody is working well, =t it could be
   darker.  We will be increasing the Ab incubation time to se=f that
   will help.

As  for  the Pan Keratin, we cannot get it = work at all. We use
   Protease  2  on  our Ultra stainer for FFPE tissues in=stology, but
   this  stain in MOHS is placed on fresh, unfixed tissue... by=anual
   drop  method.  Any  time we've tried to use an enzyme retrieval, th=
   tissue looks eaten up... even if we incubate if for a minute.

   =ASince  the tissue is not fixed, I figured that no retrieval step
   would  be=eded, but the Pan Keratin refuses to work with or without
   retrieval.
   =A

For  those of you in MOHS labs that are using a manual staining me   thod  for  Melan A and Pan Keratin, would you be willing to share your
   protoc= with us?

Thanks so much!!

Karen

K=en L. Bauer HTL/HT (ASCP) | Histology Supervisor | Pathology |
   MOHS   Lab   S=ervisor  |  Dermatology  |  Phone:  715-838-3205  |
   [9]bauer.karen <@t> mayo.edu<[10]mailto:bauer.karen <@t> mayo.edu> | Mayo Clinic
   Health  System  |  122=hipple  Street  |  Eau Claire, WI 54702 |
   [11]mayoclinichealthsystem.org<http://www.[12]mayoclinichealthsystem.o
   rg/>

   =A

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References

   1. 3D"mailto:barbara.tibbs <@t> accurated   2. ="mailto:Bauer.Karen <@t> mayo.edu"
   3. 3D"mailto:histonet <@t> lists.utsouthwestern.edu   4. 3D"mailto:barbara.tibbs <@t> accuratediagnosticlabs.c   5. 3D"mailto:histonet-bounces <@t> lists.utsouthwestern.edu   6. 3D"mailto:histo   7. 3D"mailto:Bauer.Karen   8. 3D"mailto:histonet <@t> lists.utsouthwestern.edu"
   9. 3D"mailto:bauer.kar  10. 3D"mailto:bauer.karen <@t> mayo  11. 3D"http://mayoclinichealt=
  12. 3D"http:/  13. 3D"mailto:Histonet <@t> lists.utsouthwestern.edu"
  14. 3D"http://lists.utsouthweste=
  15. 3D"mailto:Histonet <@t> lists.u  16. file://localhost/tmp/3D"h_______________________________________________
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