[Histonet] PAP stain troubleshooting

[Terri Braud]

Hi Charles - Not to worry.  Many of us Histo folks don't have a Cytologist to help.  Beth is spot on in her advice.  I just wanted to add  Orange G should appear yellow to orange; 15 sec to 1 minute is the usual range of staining times. EA is often problematic because of fundamental limitations in its chemical composition. Ideally, one should see clearcut hues of green and red in separate cells. Staining times less than about 3 minutes usually favor the uptake of eosin, with eosin and light green often occupying different areas of the same cells. Most EA formulations perform optimally in the 6-8 minute range. Note particularly that the OG and EA staining times are interdependent: relatively too much time in OG will overload cells with orange G and block the subsequent uptake of eosin.  Make sure you record your lots of stain as they are changed out, and try using a self-made buccal smear to check new lots of of the stain components before they are put into use.  Then, if you see a test problem, you can repeat the buccal smear and compare to the original.  It may help you to pinpoint the problem.
Best of luck - Terri
Terri L. Braud, HT(ASCP)
Anatomic Pathology Supervisor
Laboratory
Holy Redeemer Hospital
1648 Huntingdon Pike
Meadowbrook, PA 19046
ph: 215-938-3689
fax: 215-938-3874

   9. PAP staining quality (Beth Cox)
Message: 9
Date: Thu, 4 Feb 2016 18:06:53 -0500
From: Beth Cox <bethcoxx at gmail.com>
Subject: [Histonet] PAP staining quality
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
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Message: 3 Date: Thu, 4 Feb 2016 09:50:18 -0500
From: Charles Riley <criley at dpspa.com>
Subject: [Histonet] PAP stain quality
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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