RE: staining JB-4 sections

From:Tony Henwood

I have used Lees stain on GMA sections and it is very good as a rapid stain (like the tol blue stain for epoxy resins). We found that for a H&E to work well on GMA sections, especially if the tissue was fixed in 10%NBF, then some modification was needed. The technique is:


1. Celestine Blue - Iron Alum.

2. Harris's Haematoxylin .

3. Buffer for Eosin Y/Phloxine B.

        Solution A     2.875ml Glacial Acetic acid in 500ml distilled water.

        Solution B     4.1g Sodium Acetate in 500ml distilled water.

    150 ml Solution A + 350 ml Solution B. pH to 4.85.

4. Eosin Y/Phloxine B.

        Eosin Y (CI 45380) 5g.

        Phloxine B (CI 45410) 0.5g.

        Buffer 500ml.

        Add 2 cystals Thymol.



1. Hydrate sections.

2. Stain in Celestine Blue 15 minutes.

3. Wash in running water.

4. Stain in Harris's Hx. 15 minutes.

5. Wash, differentiate and blue till surrounding plastic is clear.

6. Stain in buffered Eosin for 6 minutes.

7. Wash lightly in running water.

8. Differentiate in alcohol till surrounding plastic is clear.

9. Clear in Xylene and mount in Eukitt.



Castro,M.D.,(1985) "A Haematoxylin - Eosin Phloxine Stain for tissues embedded in Glycol Methacrylate" J. Histotechn 8(1); 23-24.

Green,G.H.,Kurnein,F., (1981) "Glycol Methacrylate embedding in General Histopathology " ACP Broadsheet 97.


Tony Henwood JP, BappSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager
The Children's Hospital at  Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: (02) 9845 3306
Fax: (02) 9845 3318

-----Original Message-----
From: MaryLou []
Sent: Friday, 31 January 2003 6:00
Subject: Fwd: staining JB-4 sections

Techniques in Plastic for Light Microscopy
National Society for Histotechnology - Region IV Symposium
June 25, 1983   Ann Arbor, Michigan
Alexandra N. Brady, M.S., MT/HTL (ASCP)
Robert Schoonhoven, HTL (ASCP)

I like the Lee's Methylene Blue -Basic Fuchsin for an H&E look-a-like.
Date: Thu, 30 Jan 2003 16:01:38 +0100
X-PH: V4.1@mailhub3
From: =?iso-8859-2?Q?Michaela_Fr=FDzkov=E1?= <>
Subject: staining JB-4 sections
To: histonet <>

Dear Colleagues,
I need an advice about staining sections embedded in JB-4 Plus. I embed in
this medium and now I need stain my sections. I wanted to use regular
haematoxylin-eosin staining, but it doesn#180#t work. Do you know something
about that? Can you recommend me some other staining?

Thank you, very much.
Michaela Fryzkova

Department of Parasitology
Faculty of Science
Charles University
Czech Republic

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