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This is very interesting. What kind of oil did you use?
Joel
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> I would have to disagree with this.
>
> I did alot of DiI labeling in fly embryos and wsa
> labeling neurons but found that in live developing
> tissue, DiI very nicely labeled membranes. Glial
> cells for example, were beautifully labeled, as were
> fine neurite processes. I suppose it was possible
> that I was seeing cytoplasmic staining, but I really
> don't think so.
>
> Anyway, I found that the molecular probes solutions
> were usually aqueous (or alcohols). I am wondering if
> you suspended your DiI in an oil solution (which is
> what i did), whether or not you wouldn't be able to
> get good membrane staining.
>
> Alice Schmid
>
> --- "Dr. Andrea J. Elberger" <[log in to unmask]>
> wrote:
>
> > Search the CONFOCAL archive at
> >
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> > WINNOK - Your outcome using DiI was due to the fact
> > that you used it in
> > live cells.
> >
> > Even in neurons, when DiI is applied to live cells
> > it is endocytosed and
> > transported in the cytoplasm; in the case of
> > neurons, it is transported
> > retrogradely to the cell body.
> >
> > It is only when you use the DiI in aldehyde-fixed
> > tissue that you see it
> > as a lipophilic dye that passively diffuses within
> > the lipid bilayer so
> > that the entire cell membrane is labeled.
> >
> > Sorry, but DiI can't give you the results that you
> > want in these
> > conditions. You are right to seek another live
> > counterstain.
> >
> > ANDREA ELBERGER
> >
> >
> > winnok wrote:
> >
> > > Search the CONFOCAL archive at
> > >
> >
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> > >
> > > Dear all,
> > >
> > > I try to use DiI-solution (molecular probes) as a
> > counterstain for the
> > > membranes of endothelial cells but usually it is
> > visible as vesicles
> > > probably due to endocytosis.
> > > I have tried several protocols, including the one
> > delivered by
> > > molecular probes. I also tried to do the labeling
> > at 4°C to prevent
> > > endocytosis but without result, still the stain
> > was visible as
> > > vesicles. I searched the list but most
> > discussions about DiI concern
> > > neurons or blood vessels. Does anybody have
> > experience with labeling
> > > endothelial cells with DiI? Or does somebody know
> > of a good non toxic
> > > red fluorescent live counterstain for the nuclear
> > membrane or nucleus,
> > > better than DiI?
> > > Many thanks in advance,
> > >
> > >
> > > Winnok De Vos (ir.)
> > > Academic Assistant
> > > ________________________________________________
> > >
> > > Faculty of bioscience-engineering, UGent
> > > Department molecular biotechnology
> > > Coupure links 653 (R224)
> > > 9000 Gent
> > > Belgium
> > >
> > > tel nr. 09/264.59.71
> > > fax nr. 09/264.62.19
> >
> >
> > --
> > Dr. Andrea J. Elberger
> > Professor, Anatomy and Neurobiology
> > Director, Confocal Laser Scanning Microscope
> > Facility
> > The University of Tennessee, Memphis
> > 855 Monroe Avenue
> > Memphis, TN 38163 U.S.A.
> > tel: 901-448-4101
> > FAX: 901-448-7193
> > <mailto:[log in to unmask]>
> >
Joel B. Sheffield, Ph.D.
Biology Department, Temple University
1900 North 12th Street
Philadelphia, PA 19122
[log in to unmask]
(215) 204 8839, fax (215) 204 0486
http://astro.temple.edu/~jbs
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