[Histonet] H&E stain problems

[Gary Gill]

Other details being equal, hand staining will beat machine staining when
slides are dipped in rinses repeatedly.  Sakura DRS-2000 has dip
("agitation") feature.  Other machines?

Gary Gill

-----Original Message-----
From: Connie McManus [mailto:convmcm <@t> cc.usu.edu] 
Sent: Tuesday, May 18, 2004 9:09 AM
To: 'Petia P Stefanova'; 'Megan Kear'; histonet <@t> lists.utsouthwestern.edu
Cc: histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] H&E stain problems


We use almost the exact same protocol... we use Surgipath Harris, but we
prepare eosin in-house.  One thing I am amazed at in this protocol is the
length of time in the acid alcohol.  Do you use an autostainer?  We have a
Leica.  The time is set to 1 second in the acid ETOH and
sometimes the sections are almost too differentiated.   I can't imagine
6 seconds in the acid ETOH!!  Even when I do H&E manually, I dip the slides
in and quickly put them in running water.  I must have a very strong
solution, I guess. Hmmmm. Interesting.  In truth, I prefer my hand stained
sections better than when they're stained automatically.

Just wondering and blabbering (hey, it's Tuesday, what do you expect??)

Connie McManus
Utah Veterinary Diagnostics Laboratory
Utah State University
Logan, UT
Phone:  435/797-1891
fax: 435/797-2805


-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Petia P
Stefanova
Sent: Monday, May 17, 2004 6:45 AM
To: Megan Kear; histonet <@t> lists.utsouthwestern.edu
Subject: Re: [Histonet] H&E stain problems

Hi,

I use Harris's hematoxylin which is also regressive and purchase my
hematoxylin and eosin /alcohol-based/ from www.surgipath.com. I get very
good H&E staining with this protocol.

REAGENT                       TIME
Xylene                                3 min.
Xylene                                3 min.
Abs. alc.                              2 min.
Abs. alc.                              2 min
95% alc.                              2 min
80% alc.                              2 min
Wash /tap water/                  30 sec.
Hematoxylin                          8 min.
Wash /tap water/                  2 min.
Acid alcohol                          6 sec.
Wash /tap water/                  2 min.
Wash /tap water/                  5 min
80% alc.                              30 sec.
Eosin                                    15 sec.
95% alc.                              10 sec.
Abs. alc.                              30 sec.
Abs. alc.                              30 sec.
Xylene                                1 min.
Xylene                                1 min.
Xylene                                Exit

Hope it helps!
Petia


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