RE: Cartilage, Alginate, and Differential Staining

From:"Smith, Allen"

Takeuchi's acriflavine (Stain Technology 37:105-109; 1962) usually
distinguishes between sulfated polysaccharides and uronic acids.
Spicer & Meyer's aldehyde fuchsin-alcian blue sequence (Am.J.Clin.Pathol.
33:453-460; 1960) almost always distinguishes between sulfated
polysaccharides and uronic acids.

Allen A. Smith, Ph.D.
Barry University
School of Graduate Medical Sciences
    Podiatric Medicine and Surgery
Miami Shores, Florida  33161-6695 

-----Original Message-----
From: Tracey Couse []
Sent: Friday, April 19, 2002 10:05 AM
To: HistoNet Server
Subject: Cartilage, Alginate, and Differential Staining

Fellow Histolanders,

I was hoping someone could provide some insight to my present dilemma. Let 
me provide some background. I have been given some alginate gels seeded 
with chondrocytes. They were formalin fixed and paraffin embedded.  I have 
been asked to stain for cartilage. Initially this sounds easy, but the 
problem is that the alginate gel tends to stain with similar properties to 
that of the cartilage of interest. It becomes very difficult to 
differentiate between the gel and the cartilage. There are some differences 
in staining intensity, but the gel is not uniform in density so it too 
stains with different intensities masking the cartilage.

I have learned that the alginate gels are comprised of linear polymers of 
guluronate and mannuronate. I would have thought that because the 
glycosaminoglycans (gags) contain sulfates, that I would have been able to 
use alcian blue to differentiate between the sulfates of the gags and 
carboxylates of the alginate. This has not been the case. To date, I have 
tried staining with safranin-o, toluidine blue, and alcian blue at pH 1 and 
pH 0.5. Again, the results have been that the alginate gel takes on roughly 
the same shade of stain as the cartilage (I am using articular cartilage 
for a positive control).

Any suggestions, tips, ideas, corrections, and/or knowledge would be 
greatly appreciated.  I am new to the cartilage and bone field so any 
insight from the HistoNet would be invaluable to me.

Gratefully yours,

Tracey Couse
Laboratory Coordinator/Histologist
Georgia Tech/Emory Center for the
    Engineering of Living Tissues
Georgia Institute of Technology
Phone: 404.385.2611
Fax: 404.894.2291

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