[Histonet] alcian blue_ precipitation

[raghul]

dear histonetters

When i try to dissolve the alcian blue (sigma) powder in acidified (ph2.5/ph1.0) dd water the stain isnt getting dissolved. When i try 1:1 alchol:distilled water adjusted to required pH its dissolving but isnt staining any of mucin cells in positive control. please help

raghul
Medbiotech res. limited
chennai, India


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[Kemlo Rogerson]

When i try to dissolve the alcian blue (sigma) powder in acidified
(ph2.5/ph1.0) dd water the stain isnt getting dissolved. When i try 1:1
alchol:distilled water adjusted to required pH its dissolving but isnt
staining any of mucin cells in positive control. please help

Raghul



Logically if 'Alcian Blue' is not dissolving in water but does dissolve
in 1:1 water:alcohol then it is the alcohol that it is dissolving in and
I don't know if it works non polar liquids, but then a mixture would be
polar wouldn't it? (I dunno, confused).



Alcian Blue I think is usually used as an aqueous solution of 1% at
various PHs. The only thing I wonder is that Alcian Blue 8GX is the
usual dye used and Alcian Blue 8XM is less soluble and is used in lower
concentrations. I gather the composition of Alcian Blue 8GX has, over
the years, been changed to make it more soluble. These are basic dyes
with a phthalocynain nucleus and basic side chain; logically you either
been sold the 'wrong stuff' (8XM) or you have a poorly soluble variant.



Send it back, get some more.



Kemlo Rogerson

Pathology Manager

DD 01934 647057 or extension 3311

Mob [phone]; Pager 07659 597107;



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[b***@cogeco.ca]

Raghul,

try dissolving the Alcian blue powder in distilled water first, then adjust the pH with acid. You should have no problem
with pH 2.5, but some lot #s of dye will precipitate at pH 1.0. If this happens contact me again.


Bryan

dear histonetters
When i try to dissolve the alcian blue (sigma) powder in acidified (ph2.5/ph1.0) dd water the stain isnt getting
dissolved. When i try 1:1 alchol:distilled water adjusted to required pH its dissolving but isnt staining any of mucin
cells in positive control. please help
raghul
Medbiotech res. limited
chennai, India
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[Kellar, Eric C]

The dye required is Alcian Blue 8G. Alcian Blue 5G and 7G are similar, but carry fewer of the cationic substituents and are less water soluble. Alcian Blue 8GX (X for extra) has a higher dye content than does 8GS (S for standard). The stability of the dye is variable. Depending on the age of your dye lot, it could take approximately 1 hour to dissolve Alcian Blue 8GX into aqueous solution with a magnetic stirrer, then adjust the pH.

Eric C. Kellar
Quest Diagnostics
Histology Laboratory Supervisor
South Florida



-----Original Message-----
From: histonet-***@lists.utsouthwestern.edu [mailto:histonet-***@lists.utsouthwestern.edu] On Behalf Of raghul
Sent: Tuesday, April 17, 2007 9:22 AM
To: ***@lists.utsouthwestern.edu
Subject: [Histonet] alcian blue_ precipitation

dear histonetters

When i try to dissolve the alcian blue (sigma) powder in acidified (ph2.5/ph1.0) dd water the stain isnt getting dissolved. When i try 1:1 alchol:distilled water adjusted to required pH its dissolving but isnt staining any of mucin cells in positive control. please help

raghul
Medbiotech res. limited
chennai, India


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[Gayle Callis]

Our pH 2.5 alcian blue was dissolved in 100 ml of 3% glacial acetic acid,
and pH 1 alcian blue was dissolved in 0.1N HCL.

Your alcian blue may be old? Or not biological stain commission
certified. We also store our alcian blue in the refrigerator, per
recommendation of R.W. Mowry of McManus and Mowry fame, his lecture at NSH
some years ago.

Post by raghul
dear histonetters
When i try to dissolve the alcian blue (sigma) powder in acidified
(ph2.5/ph1.0) dd water the stain isnt getting dissolved. When i try 1:1
alchol:distilled water adjusted to required pH its dissolving but isnt
staining any of mucin cells in positive control. please help
raghul
Medbiotech res. limited
chennai, India
---------------------------------
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Gayle Callis
MT,HT,HTL(ASCP)
Research Histopathology Supervisor
Veterinary Molecular Biology
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610

[Akemi Allison-Tacha]

This is what I do with my Alcian Blue. My stabalizer
is missing from this protocol because it is
proprietary. By the way, Anatech has the best Alcian
Blue on the market! The most dye content!!!

1. Label the appropriately sized container with
description, lot number, storage conditions,
expiration date, prepared by and date. Add a large
stir bar to the container.

2. Measure the 3% acetic acid solution determined in
the Calculation section of Manufacturing Document.
Transfer the 3% Acetic Acid to the labeled container,
and begin stirring on a large stir plate. Add the
Alcian Blue and continue mixing solution.

3. Mix thoroughly for 30±5 minutes or until all the
chemicals and reagents have dissolved. Visually
inspect solution to ensure adequate mixing.

4. Measure the pH of the solution using a pH meter
calibrated at pH 1 and pH 4. The pH must be 2.5 ±
0.1. If necessary, adjust pH with NaOH of HCL.

5. The reagent is stable for 12 months at room
temperature. Store the reagent at room temperature in
an opaque container.

6. Test for quality control.

Good Luck,
Akemi Allison-Tacha

Post by raghul
dear histonetters
When i try to dissolve the alcian blue (sigma)
powder in acidified (ph2.5/ph1.0) dd water the stain
isnt getting dissolved. When i try 1:1
alchol:distilled water adjusted to required pH its
dissolving but isnt staining any of mucin cells in
positive control. please help
raghul
Medbiotech res. limited
chennai, India
---------------------------------
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Yahoo! Messenger
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Histonet mailing list

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[Tony Henwood]

Dissolve the dye in water first and then add diluted acid in water.
Eg for 1% alcian blue in 2.5% acetic acid:
1g dye in 50ml distilled water.
2.5ml acetic acid in 47.5ml water.
When the dye has dissolved, mix the two solutions

Regards

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: [phone]
Fax: [phone]




-----Original Message-----
From: histonet-***@lists.utsouthwestern.edu
[mailto:histonet-***@lists.utsouthwestern.edu] On Behalf Of raghul
Sent: Tuesday, 17 April 2007 11:22 PM
To: ***@lists.utsouthwestern.edu
Subject: [Histonet] alcian blue_ precipitation


dear histonetters

When i try to dissolve the alcian blue (sigma) powder in acidified
(ph2.5/ph1.0) dd water the stain isnt getting dissolved. When i try 1:1
alchol:distilled water adjusted to required pH its dissolving but isnt
staining any of mucin cells in positive control. please help

raghul
Medbiotech res. limited
chennai, India


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***@lists.utsouthwestern.edu
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[Tony Henwood]

Alcian blue tetrakis (methylpyridinium) chloride (Sigma, Cat. No. A4045,
90% dye content) is a better dye to use. See "Improved demonstration of
mast cells using alcian blue tetrakis (methylpyridium) chloride"
Biotechnic & Histochemistry 77(2):93-94.(2002)

Regards

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: [phone]
Fax: [phone]




-----Original Message-----
From: histonet-***@lists.utsouthwestern.edu
[mailto:histonet-***@lists.utsouthwestern.edu] On Behalf Of
***@cogeco.ca
Sent: Wednesday, 18 April 2007 12:09 AM
To: raghul; ***@lists.utsouthwestern.edu
Subject: re: [Histonet] alcian blue_ precipitation

Raghul,

try dissolving the Alcian blue powder in distilled water first, then
adjust the pH with acid. You should have no problem with pH 2.5, but
some lot #s of dye will precipitate at pH 1.0. If this happens contact
me again.


Bryan

dear histonetters
When i try to dissolve the alcian blue (sigma) powder in acidified
(ph2.5/ph1.0) dd water the stain isnt getting dissolved. When i try
1:1 alchol:distilled water adjusted to required pH its dissolving but
isnt staining any of mucin cells in positive control. please help
raghul
Medbiotech res. limited
chennai, India
---------------------------------
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_______________________________________________
Histonet mailing list
http://lists.utsouthwestern.edu/mailman/listinfo/histonet

_______________________________________________
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This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender.

Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead

This note also confirms that this email message has been
virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses.
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