[Histonet] Safranin O cartilage staining

Elizabeth Chlipala liz <@t> premierlab.com
Wed May 22 09:14:26 CDT 2013


Safranin O can be a bit tricky at times.   There are a few tips that we have used through the years.

1.  Fresh reagents are key.
2.  For proteoglycan staining you can cut the sections a bit thicker and get better staining we typically cut the joint sections for Saf O at 6 to 7 microns in thickness, there are papers out there that recommend up to 8 microns in thickness.
3.  Limit excess time in decal for some reason this particular stain may not work as well if the samples are in decal for an extended period of time, we have not seen this with toluidine blue which is another stain for proteoglycan.
4.  We increase times in the safranin O reagent on occasion for the murine joints.

Good Luck


Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Laboratory Manager
Premier Laboratory, LLC
PO Box 18592
Boulder, CO 80308
Work (303) 682-3949
Fax (303) 682-9060
Cell (303) 881-0763
liz <@t> premierlab.com

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-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Brett Tonkin
Sent: Tuesday, May 21, 2013 11:08 PM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Safranin O cartilage staining


We're having trouble staining murine articular cartilage with Safranin O. We first started using this stain last year and it was working nicely, with strong staining of both the articular cartilage and growth plate. After a while, the stain stopped working, with staining only visible in the growth plate. We replaced all solutions (including c/stain of fast green) and the staining worked. Yet again, it has stopped working, and this is only the second time the solutions have been used.

Has anyone come across this before?

Any help or advice would be greatly appreciated!

Brett Tonkin

Research Assistant
Arthritis Research Laboratory
Bone Cell Biology and Disease Unit
St. Vincent's Institute
Fitzroy, Victoria

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