[Histonet] ihc slides

Greg Dobbin gvdobbin <@t> ihis.org
Mon Jan 24 10:31:04 CST 2011

Hematoxylin will inactivate the peroxidase (link) in the tissues. You must go back and re-do the link step and hopefully the antigenic sites will still accept more streptavidin (or similar) conjugated antibody.
Greg Dobbin, R.T.
Chief Technologist, Anatomic Pathology
Dept. of Laboratory Medicine,
Queen Elizabeth Hospital,
P.O. Box 6600
Charlottetown, PE    C1A 8T5
Phone: (902) 894-2337
Fax: (902) 894-2385
"I find that the harder I work, the 
more luck I seem to have."
- Thomas Jefferson

>>> "Cynthia Pyse" <cpyse <@t> x-celllab.com> 1/24/2011 8:57 AM >>>
Happy Monday Histonetters

I need to pick the brains of the IHC experts out there in histoland. I run
IHC on the Dako autostainer. On one of our runs there was not enough DAB to
dispense on all the slides. My tech dehydrated the slides and cover slipped
them. We printed the stain log on the run and confirmed that the DAB was not
dispensed on three slides. The cover slips were removed and the slide were
soaked in xylene to remove the cover slipping media. The slides were
rehydrated and DAB was applied. There was still no reaction after 10
minutes, The DAB was reapplied for up to 30 minutes with no reaction. Can
anyone out there give me a reason that the slides would not react after the
reapplication of the DAB. I have done this before when the DAB solution was
short on the run and it worked. I can't figure out why this didn't react. I
called tech service at Dako and talked to the supervisor, she agrees with me
that the reapplication should have worked. I checked the run log and the
only missing step is the DAB. The antibodies with the missing DAB have their
own controls on the slides. Any thought out there as to what the problem is.
I have had the PM done on the Dako which should resolve the problem
according to Dako. There has not been a problem since. This mystery will
drive me crazy until I can figure out why. I could use help. Thanks in
advance, I appreciate any input.


Cindy Pyse, CLT, HT (ASCP)

Laboratory/Histology Supervisor

X-Cell Laboratories


e-mail cpyse <@t> x-celllab.com

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