Epithelial Tissue Fixation for Fluorescent Imaging

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Epithelial Tissue Fixation for Fluorescent Imaging


I am looking to fix epithelial tissue (multiple layers of epithelial cells) grown on a 0.4um pore poly-carbonate membrane cell culture insert  (from NUNC).

what is the best way to fix the tissue?

1- para- formaldehyde? I think its for cells not tissue?

2- I tried -20C methanol for 18 hrs, followed by wash with acetone  and then incubation with primary and secondary antibody and post fixed with ethanol. It didn't give great results. was this too harsh? Note that the poly carbonate membrane is still entact. I am not looking to see the cross section but a tight junction protien occludin between epithelial cells.
Any good protocol for fixing tissue for such a study?