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Dear confocalists,
change of the medium may be a way to get rid of autofluorescence. But
consider that the composition of the culture medium may substantially
influence the physiological state of the cells (e.g. protein expression). I
was puzzled with such an observation several years ago when studying
induction of vimentin expression in mouse myeloma cells (Giese G. and
Traub, P., Eur. J. Cell Biol. 47, 291-299 (1988)
Serum itself is a "multivariate standard" (Homm et al.: Fetal Bovine Serum:
A multivariate Standard. Proc. Soc. Exp. Biol. Med. 149 (1975) 344-347).
Sometimes using a new lot of your standard serum may help (introducing less
possible side effects of medium change). Ask your cell culture facility to
order and test new lots of serum.
Some serum-free media may also exert special effects.
Therefore: check possible side effects of changing the growth medium.
Guenter
At 10:27 21.10.02, you wrote:
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>Hello Amy,
>There seems to be a possibility that this is caused by substances from
>the culture medium. We found it in slime mould cells that were grown in
>a medium containing DMEM, but not in phosphate buffer. I suppose it has
>something to do with the serum component. If you are referring to fixed
>specimens it is, of course, possible that you are observing fixation -
>induced fluorescence (glutaraldehyde ?!?!). I remember having been
>annoyed by varying degrees of fluorescence in formaldehyde fixed slices
>of liver- some cells showing a lot, others only some fluorescence. At
>that time I assumed it was related to varying physiological functions,
>but I never followed it up. We found no way to come around it, like
>keeping the cells in serum deprived medium before fixation (this might
>have helped if fluorescent substances in the endosome-lysosome
>compartment would be digested)or thorough treatment with Borohydride
>(which would have quenched the aldehyde -induced fluorescence). I should
>be grateful for any info or suggestion myself.
>Wessel
>Wessel de Priester
>Leiden University
>IBL - EMCA
>
>-----Original Message-----
>From: Shen, Amy C [mailto:[log in to unmask]]
>Sent: vrijdag 18 oktober 2002 22:41
>To: [log in to unmask]
>Subject: Autofluorescent hepatocytes?
>
>
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> > Hello, We observed very bright and definite fluorescence (red
>and
> > green) in our primary cultured and cryo-preserved hepatocytes. I
>wonder
> > if anybody has observed that or not. The cells were grown on Collagen
> > I-coated chamber slides. Can someone enlighten me, and also is there
> > anything one can go around this? Thanks. Amy Shen.
>
>
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------------------------------------------
Dr. Guenter Giese
Light Microscopy Facility
Dept. of Biomedical Optics, MPI fuer Medizinische Forschung
Jahnstr. 29, D-69120 Heidelberg, Germany
Phone (+49) 6221-486-360 (Fax: -325)
e-mail: [log in to unmask]
http://sun0.mpimf-heidelberg.mpg.de/~ggiese/lightmicro/index.html
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