H&E for 5um MMA sections

From:Cathy Mayton <Cathy@wasatchhisto.com>

Dear Vicki and Jennifer,

The following can be used for the staining of 5 micron MMA sections.  This
was taken from a workshop I presented in 1989 at NSH.  If you are
interested, you may visit my website at www.wasatchhisto.com  There I have
listed all of my publications most of which are for ground histology. 

H&E Staining of 5 micron MMA Sections

Deplastify  sections first by placing slides in a tightly capped coplin jar
containing xylene.  Place in a 60 degree C oven overnight.  Remove from the
oven and allow xylene to come to room temperature.  Hydrate slides to tap
water.  Rinse in distilled water and stain as follows:

1.  Hydrate slides to tap water, then rinse in distilled water.
2.  Stain 6 minutes in hematoxylin of your choice (I used Surgipath's Gill
3.  Rinse excess stain in running tap water.
4.  Decolorize with 1-2 quick dips in 0.5% acid alcohol and rinse in tap
5.  Dip sections in ammonia water (12 drops in 250 ml distilled water) and
blue nuclei in running tap water for 5 minutes.
6.  Rinse in distilled water.
7.  Dip slides in 5-10 times in 80% alcohol.
8.  Stain 1 minute in eosin of your choice (I use eosin/phloxine).
9.  Dehydrate in graded alcohols (95-100%).
10. Clear slides in xylene or xylene substitute.
11. Mount sections with mounting medium of your choice (must be compatible
with clearing reagent).

I would have to look back in my publications but I believe we first used
cellosolve (sold by Fisher) to deplastify sections for 25 minutes but then
read about Tony Villanueva using 60 degree xylene.  We stayed with the
xylene because it seemed to work better.

I hope this is helpful.  Good luck with your staining.  Please call or
write me if you think I can help you futher.

Best regards,
GLP Compliant Laboratory 

Cathy A. Mayton
Project Director
Wasatch Histo Consultants, Inc.
80 Youngberg Road
Winnemucca, NV   89445-5709
Voice:  (775) 625-4425
Fax: (775) 625-1548

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