I, too, was taught that osmium tetroxide reated with the unsaturated lipids
in the cell membrane.  I cannot see how this can be true.  The lipids are
the middle layer of the membrane.  If osmium tetroxide reacted with the
unsaturated lipids, we would see the cell membrane as a single dark gray
line in electron micrographs.  Since we see the membrane as two black lines,
osmium tetroxide must be reacting with the amino alcohols which form the two
outside layers of the membrane.
    Osmium tetroxide reacts with fat in fresh tissue by reacting with the
double bonds in oleic acid.    If one cuts thin sections for electron
microscopy, fat droplets are gray because there isn't that much oleic acid
present.  If one cuts thick sections, one sees fat droplets as black because
many layers of gray look black.

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

----- Original Message -----
From: Morken, Tim <tim9@cdc.gov>
To: <Histonet@Pathology.swmed.edu>
Sent: Thursday, May 17, 2001 1:05 PM
Subject: RE: Re Oil red staining


> Well, for whatever reason osmium will preserve large fat droplets in
tissue
> and I have used it to demonstrate lipid storage diseases in paraffin
> sections. So, I can't give you the chemistry, just the observed results.
>
> Tim
>
> -----Original Message-----
> From: Philip Oshel [mailto:peoshel@facstaff.wisc.edu]
> Sent: Thursday, May 17, 2001 11:21 AM
> To: Morken, Tim
> Cc: Histonet@Pathology.swmed.edu
> Subject: RE: Re Oil red staining
>
>
> A question: is this really the case? Osmium binds across double-bonds
> of lipids, which is why it shows and preserves membranes, but it
> binds poorly or not all at to saturated lipids. So I wouldn't expect
> OsO4 to show fatty deposits if the fats are saturated. Oil Red O and
> Sudan Black, which more mix into the lipids and don't react with
> them, would show fat deposits that OsO4 doesn't, and are better fat
> stains.
>
> Phil
>
> >Agreed, there would be no need to do special fat stains if the tissue is
> >processed with osmium. In fact, this can be done for paraffin embedding
as
> >well to show fat distribtion in some diseases with vastly better
morphology
> >and localization than frozen sections.
> >
> >Tim Morken
> >CDC, Atlanta
> >
> >-----Original Message-----
> >From: Saby, Joseph [mailto:Joseph.Saby@pfizer.com]
> >Sent: Thursday, May 17, 2001 6:15 AM
> >To: 'RichardWHorobin@aol.com'; Histonet@pathology.swmed.edu
> >Subject: RE: Re Oil red staining
> >
> >
> >Richard-
> >
> >Whenever I've worked with those plastics, there has always been a
clearing
> >stage through acetone.  Since acetone would remove all non-bound fat, Oil
> >Red O would have nothing to go into.
> >
> >When I've worked with these plastics, I also did a post-fixation in
osmium
> >tetroxide, which does a very good job of staining fats and lipid (it
turns
> >them black).  Perhaps this would work for your purposes.
> >
> >Joe
> >
> >Joseph A. Saby, BA, HT(ASCP)
> >Drug Safety Evaluation
> >Pfizer Global Research and Development
> >2800 Plymouth Road
> >Ann Arbor, MI 48105
> >Phone: (734)-622-3631
> >FAX:   (734)-622-3866
> >E-mail: joseph.saby@pfizer.com
> >
> >
> >
> >-----Original Message-----
> >From: RichardWHorobin@aol.com [mailto:RichardWHorobin@aol.com]
> >Sent: Thursday, May 17, 2001 2:49 AM
> >To: Histonet@pathology.swmed.edu
> >Subject: Re Oil red staining
> >
> >
> >Jaclynn Lett
> >wrote:
> >
> >
> >
> >Has anyone used Oil Red O to stain lipids in tissues embedded in plastics
> >(Epon or Epon-Araldite)?  If so, has this been done by staining en bloc
or
> >by staining the sections.  Sections would range from 1-4 microns in
> >thickness.
> >
> >
> >
> >A couple of people so far have commented on how hard this would be, and
our
> >experience agrees with theirs.
> >HOWEVER Jaclynn also said:
> >
> >
> >
> >
> >We would also consider tissues embedded in glycol methacrylate.  We'd
like
> >to avoid frozen sections because we'd prefer the higher level of detail
> >possible with plastic.
> >
> >
> >
> >In THAT case things look better! Would you settle for Sudan black, rather
> >than Oil red staining of the lipid? If 'yes' then there is a method -
which
> >does indeed show even tiny droplets of lipid very clearly. This was
worked
> >out by the one-time king of GMA staining Peter Gerrits, and can be found
in
> >J
> >Neurosci Methods, as follows:
> >
> >      Gerrits PO, Brekelmans-Bartels M, Mast L, 's-GrAavenmade EJ,
Horobin
> RW
> >
> >and Holstege G. (1992)..
> >      Staining myelin and myelin-like degradation products in the spinal
> >cords of chronic experimental
> >      allergic encephalomyelitis (Cr-EAE) rats using Sudan Black B
staining
> >of glycol methacrylate-embedded
> >      material.
> >      J. Neuroscience Methods. 45, 99-105
> >
> >Bye - Richard Horobin
> >
> >Institute of Biomedical & Life Sciences, University of Glasgow
> >T direct 01796-474 480 --- E  RichardWHorobin@aol.com
> >"What should we expect? Everything."
>
> --
> }}}}}}}}}}}}}}}}}{{{{{{{{{{{{{{{{{
> Philip Oshel
> Supervisor, AMFSC and BBPIC microscopy facilities
> Department of Animal Sciences
> University of Wisconsin
> 1675 Observatory Drive
> Madison,  WI  53706 - 1284
> voice: (608) 263-4162
> fax: (608) 262-5157 (dept. fax)
>
>