RE: [Histonet] automated IHC on bone sections?

From:"Elizabeth Chlipala"


I have run FFPE, formic acid decalcifed joint sections on the Dako
autostainer.  I try to stick with enzyme digestion for the joint
sections.  I have ran F4/80, IL-6 and complement C3 with success on the
autostainer.  The F4/80 and IL-6 require proteinase K digestion and that
can be done on the autostainer. I'll e-mail you off the histonet some


Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Premier Laboratory, LLC
P.O. Box 18592
Boulder, Colorado 80308
Office: (303) 735-5001
Fax: (303) 735-3540
Ship to Address:
Premier Laboratory
University of Colorado
MCDB, Room A3B40
Boulder, Colorado 80309

-----Original Message-----
[] On Behalf Of Garlits,
Sent: Thursday, January 13, 2005 3:10 PM
Subject: [Histonet] automated IHC on bone sections?


I am writing to ask if anyone out there has successfully done automated
IHC on decalcified, paraffin-embedded sections.  I have been staining
these for GFP, manually, for some time, but I have found that my IHC
results are quite variable.  I would like to work out an automated
method that might be faster and more consistent.  I have submitted
sections for automated staining at an in-house facility, but the problem
is that the actual bone part of my mouse femur sections always peels
off.  I use an enzyme-based antigen retrieval method when I do them
manually, but I understand that automated stainers typically use a
heat-based method.  I suspect that the heat-based retrieval is what
makes the bone peel off because I have tried a manual version of this
and witnessed the same result.  Do these automated stainers have the
ability to omit the heated retrieval step, or even pause it to introduce
an enzyme solution?

John Garlits
St. Jude Children's Research Hospital
Memphis, TN

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