Re: Live cell imaging prep. for yeast S. pombe

Posted by mahogny on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Live-cell-imaging-prep-for-yeast-S-pombe-tp2955849p2955915.html

Graham Wright wrote:

> Hello,
>
> We are doing some live cell imaging of Schizosaccharomyces pombe
> (fission yeast). Usually the cells are mounted on an agar plug (agar &
> YES media) on a concave slide, covered with a coverslip and sealed.
> This works very well for confocal microscopy, but recently upon trying
> widefield microscopy we have noticed a significant background
> fluorescence, presumably from the agar. Simply mounting the cells in
> liquid media on a normal glass slide significantly removes the
> background problem, but creates another - the cells do not adhere to
> the glass and so make long term timelapse imaging very difficult.
>
> So, can anyone suggest ways of encouraging the cells to adhere that
> work well for yeast, particularly S. pombe? Alternatively any
> suggestions of agar, or other gelling agents that do not fluoresce
> would be appreciated.
>
> Thanks in advance for your help,
> Graham
>  

for c.elegans embryos we use polylysine to glue them down. to make
sure they aren't crushed, we add ~10 50um beads.

polylysine seems to work best if you add a droplet and then lets it dry
on a heat block, probably due to some crystallization.

/Johan

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Johan Henriksson
MSc Engineering
PhD student, Karolinska Institutet
http://mahogny.areta.org http://www.endrov.net