we use a non IHC Azure B stain to differentiate collagens
We use a simple Azure B stain for this.
0.2% Azure ph 1 for cartilage
0.2% Azure ph 4 for bone and cartilage
0.2% Azure ph 7 for bone, cartilage, collagen, everything
This is a metacromatic stain and will stain the different components different
colors (pink to blue)
Deparaffinze to water
stain sections in azure for 5 min.
rinse in deionized running water till clear
air dry (dehydrating thru alcohols and xylene will remove the stain)
coverslipp in permount
"Smith, Allen" wrote:
> 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 [mailto:email@example.com]
> 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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