[Histonet] Victoria Blue stain

[LuAnn Anderson]

VICTORIA BLUE - NUCLEAR FAST RED FOR HBSAg

PRINCIPLE
To demonstrate copper associated protein, elastic 
and Hepatitis B surface antigen (HBSAg) in paraffin wax sections.

REFERENCE
. AFIP laboratory methods in Histotechnology p210 
Tanaka method Ref:Acta Pathol Jpn 1981;31:93

SPECIMEN
10% neutral buffered formalin. Standard paraffin wax sections.

CONTROL
Known case of primary biliary cirrhosis.

REAGENTS
(A) VICTORIA BLUE
Distilled water 200 mL Dextrine 0.5 g Victoria 
Blue 2 g Resorcinol 4 g Gradually warm up the 
mixed solution of all the above until it boils. 
Remove form heat then gradually add 25mL of 
boiling 29% ferric chloride solution (12mL 
commercial 60% ferric chloride solution plus 13mL 
distilled water) and boil the solution for a 
further 3 minutes. Cool. After cooling filter it. 
Dry the filtrate on the filter paper in a 56øC 
oven until completely dry. Dissolve the dried 
filtrate in 400mL of 70% alcohol. Finally add 4mL 
concentrated hydrochloric acid and 6g phenol. It 
is best to leave this solution to mature for at least two weeks prior to use.
(B) NUCLEAR FAST RED
Dissolve 0.5g nuclear fast red in 500mL of warmed 
5% aluminium sulphate. Filter when cool.
(C) 4% AQUEOUS SODIUM METABISULPHITE

PROCEDURE
1. Deparaffinize and hydrate to distilled water.
2. Treat with acidified potassium permanganate 
(as for Gordon and Sweet's Reticular fibres method 2.4) for 5 minutes.
3. Treat with 4% sodium bisulphite for 1 minute.
4. Wash in running tap water.
5. Wash well with 70% alcohol.
6. Stain in the Victoria Blue solution, in a 
Coplin jar, for a minimum of 4 hours (preferably overnight).
7. Wash well with 70% alcohol for approx 1 
minute. This is the differentiation step - ensure 
the background of the section is clear.
8. Wash in running tap water for 1 minute.
9. Stain with nuclear fast red solution for 5 minutes.
10. Wash in running water for 2 minutes.
11. Dehydrate, clear and mount.

RESULTS
HBsAg, elastic fibres, copper associated protein blue
All other tissue components pink/red

HEALTH AND SAFETY
Potassium permanganate Harmful by ingestion. 
Irritating to eyes and skin Ferric chloride 
Irritant. Hydrochloric acid Vesicant. Causes 
burns. Phenol Harmful by ingestion. Irritating to eyes and skin.








At 12:30 PM 11/4/2009, Justin Peters wrote:
>One of my pathologists has requested that we start offering a Victoria
>Blue stain for liver specimens.  I am unfamiliar with this stain and was
>wondering if anyone had a protocol for this that works well?  Thanks in
>advance for any help :-)
>
>
>
>Justin Peters, HTL (ASCP)
>
>IHC Supervisor
>
>Bostwick Laboratories(tm)
>For Absolute Confidence(r)
>
>4355 Innslake Drive
>Glen Allen, Virginia 23060
>Phone:    (804) 967-9225 ext. 1831
>Cell:        (804) 822-6084
>Email: jpeters <@t> bostwicklaboratories.com
><mailto:jpeters <@t> bostwicklaboratories.com>
>
>
>
>_______________________________________________
>Histonet mailing list
>Histonet <@t> lists.utsouthwestern.edu
>http://lists.utsouthwestern.edu/mailman/listinfo/histonet