Frozens, fix in paraformadehyde, or look at them unfixed, NBF has also been
used. Can the cell monolayers be examined directly, WITHOUT all the extras.
 We look at monolayers, grown on chamber slides, pull off the gaskets and
coverslip, examine the monolayers without sectioning.   Clontech has a
Living Colours Manual giving hints on looking at GFP, etc and info on
autofluorescence.

Cells can be released from flasks, rinsed with PBS 3 X, spinning each time
to pellet cells, suspend in OCT gently, snap freeze and section.  Do this
in a 50 ml conical, pop the little block out by warming the outside a
tidge, hit the end to release block, and mount on a chuck.  Narrow conical
tubes are difficult for block removal.





 



At 12:12 PM 2/2/01 -0500, you wrote:
>Hello all,
>
>Is there a way to prepare for sectioning cell monolayers which express GFP
>without losing the fluorescence?  We want to do confocal microscopy on
>semi-thin sections. We could do paraffin, frozen, or any type of resin
>embedded samples.
>
>Any helpful hints out there?
>
>Thanks!
>
>Dotty Sorenson
>Microscopy and Image Analysis Laboratory
>Department of Cell and Developmental Biology
>University of Michigan Medical School
>Ann Arbor, Michigan
>(734)763-1170
>FAX (734)763-1166
>dsoren@umich.edu
>
>
>
>
Gayle Callis
Veterinary Molecular Biology
Montana State University - Bozeman
Bozeman MT 59717-3610

406 994-6367
404 994-4303 (FAX)