[Histonet] AutoStainer XL Tweaking

[Bartlett, Jeanine (CDC/CCID/NCZVED)]

Which type of hematoxylin, etc. are you using? (I know you said Sigma,
but do they have more than one formula?) Are you using their acid
alcohol or are you making your own?


Jeanine Bartlett, BS, HT(ASCP)QIHC
Centers for Disease Control and Prevention
Infectious Diseases Pathology Branch
1600 Clifton Road, MS/G-32
18/SB-114
Atlanta, GA  30333
(404) 639-3590 
jeanine.bartlett <@t> cdc.hhs.gov



-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Breeden,
Sara
Sent: Tuesday, October 16, 2007 8:04 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] AutoStainer XL Tweaking

I'm setting up my Leica AutoStainer XL and would appreciate any help
with tweaking the times.  I based my timing for the Autostainer on my
manual stain times but staining is not crisp and looks washed out.  Docs
say eosin is fine but cellular detail is foggy.  Here are my times:

XYL - 3 at 5 min each

100% reagent - 2 at 30 seconds each

95% reagent - 1 at 30 seconds

80% reagent - 1 at 30 seconds

DI water wash - 1 minute

Hematox (currently using Sigma) - 5 minutes

DI water wash - 1 minute

Acid Alcohol (1%) - one dip

DI water wash - 1 minute

Ammonia water - 15 seconds

DI water wash - 1 minute

80% reagent - 1 at 30 seconds

95% reagent - 1 at 30 seconds

Eosin (alcoholic) - 10 seconds

95% reagent - 1 at 30 seconds

100% reagent - 2 at 1 minute each

XYL - 3 at 1 minute each

I cut my acid alcohol time down from 15 seconds to one dip but it's
still taking a lot of hematoxylin off.  Should I try 0.5% acid alcohol?
I'm at a loss so far; different hematox? More time in ammonia water?  If
you use this Leica AutoStainer XL, could you offer some Handy Hints?  I
thank you SO much!

 

Sally Breeden, HT(ASCP)

NM Dept. of Agriculture

Veterinary Diagnostic Services

PO Box 4700

Albuquerque, NM  87106

505-841-2576

 

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