[Histonet] Unsuccessful Muscle Lipid Staining

Liz Chlipala liz <@t> premierlab.com
Sun Jul 22 11:13:27 CDT 2007


We did this in the past, we post fixed in osmium, it turned out quite =
nice.  I'll send images in a different e-mail.  We fixed in formalin and =
then post fixed the muscle in osmium overnight, we were working on mouse =
tibialis anterior so we kept the entire muscle intact and processed the =
whole muscle and then cut the muscle in half prior to embedding.  We =
found that the unstained sections turned out the best for demonstration =
of the fat. I might have a material and methods some where since I think =
they published on it.  I'll send that to you with the images.


Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Premier Laboratory, LLC
P.O. Box 18592
Boulder, CO 80308
phone (303) 735-5001
fax (303) 735-3540
liz <@t> premierlab.com
Ship to Address:
Premier Laboratory, LLC
University of Colorado at Boulder
MCDB, Room A3B40
Boulder, CO 80309

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu =
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Sonny =
Sent: Sunday, July 22, 2007 3:31 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Unsuccessful Muscle Lipid Staining

Hi all,

I have been trying to stain mouse muscle tissue for intramuscular =
triglyceride droplets with Oil Red O (dissolved in triethyl phosphate) =
for some time now, and my results have been disastrous.  I've tried it =
on both fresh frozen and formaldehyde fixed tissues (and cryoprotected) =
cryosections, but almost always it seems that the lipid droplets within =
the fibers (maybe?) leak out of the cells and conglomerate on the edges =
of the tissue and within gaps between the cells.  Very few of the =
fibers, if any, display any lipid droplet staining and it is always very =
weak. The lipid droplets all conglomerate in roughly the same positions =
between serial sections as well, which leads me to believe something is =
happening during the freezing/fixation process, or the cutting in the =
cryosection.  Does anyone have any suggestion as to what I could be =
doing wrong here (i.e cutting technique, temperature, fixatives)?  Also, =
does anyone have any experience with using ORO in triethyl phosphate?

Thank you all for your time,
Sonny Duong
University of Virginia
Green Lab

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