Hi Everybody!
 
I'm a fairly new confocalist (we have recently bought a Leica TCS 4d with
inverted microscope).  A researcher here wants to localise internalised
structures in BHK cells infected with dengue virus.  He initially fixed the
cells with cold acetone for 1 minute but this caused dissolution, and hence
total loss of structure, of the membrane.  To minimise damage to the
membrane, he tried fixing in ethanol.  Unfortunately, the membrane remained
intact, so much so that the antibody-FITC conjugate couldn't penetrate.  He
is now talking about slam freezing his cells, sectioning them in a
cryoultramicrotome, followed by labelling.  This seems tedious to say the least.
 
Can someone recommend a fixation agent/schedule to give both adequate
morphology and adequate access also?
 
Many thanks,
 
Felicity Lawrence
Analytical Electron Microscopy Facility,
QUT, Brisbane.  Australia.