free-floating section staining
on 29/10/2002 11:27 AM, Greg Dobbin at dobbin@Upei.CA wrote:
> HI Melissa,
> I don't have a lot of experience with this, but what I did do for
> someone else one time, was to cut 50-80 um frozen sections and
> place them directly in a 24-well plate (each section to its own well).
> The 24-well plate was kept in the cryostat and the fixative (I forget
> now, which one they were using) was already dispensed to all the
> wells. All subsequent staing took place in the well to minimize
> tissue handling (ie the reagents were changed rather than moving
> the tissue sections).
if you do use 24-well plates, be sure to get the ones _not_ treated for
tissue culture. these are made to be 'sticky' for big molecules in order
for cells to attach to the plastic, and this property can interfere with
many staining procedures. immunolabeling is especially likely to be
affected. if you do have to use culture-treated plates, at least do a
pre-incubation step, putting reagent in the wells without tissue sections,
then using fresh reagent when the tissue samples are present. in my lab we
use Falcon 1147 polystyrene plates but have recently found a source for
polypropylene plates (Phenix) which should be even better. they're cheaper
and reusable too.
we transfer sections (from the knife, and into slide mounting buffer after
staining) with small paintbrushes, but then for most procedures the
sections stay in the same well until the staining is done. we use flamed
(smooth tip) pasteur pipettes to remove solutions and micropipettors to add
solutions to the wells.
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