I have been DAPI staining fixed worms that have been stained with X-GAL.
They have all been fixed together but seem to have different permeabilities
to DAPI (?). In one batch I can see worms that are stained throughout with
DAPI in others just the embryos inside the adult are stained and in others
I observe only the posterior half of the worm is stained with the staining
NOT being in the anterior part of the intestine nor in the anterior arm of
the gonad. In these worms though all the embryos are stained. In all
these situations the staining for b-gal is seen in both anterior and
posterior cells, so the X-GAL is not exculded. In case anyone is
wondering, I incubated them in .5ug/ml DAPI-PBS for 15 min. observed they
were not stained "evenly" and thinking it was the exposure time I just
mounted them with DAPI in the mounting medium. They have remained with
this odd staining.
I have discussed this with other non-worm people and they have suggested
freeze-thawing them a few times or warming them up. Does anyone know of
why this happens and how it can be fixed?