[Histonet] Improving morphology in cryosections of human eye

From:"Johnson, Teri"

A researcher here is trying to accomplish IHC on 10 micron sections of fresh, frozen human eye.  Essentially she uses the CryoJane tape transfer system, then air-dries the slide, fixes for 15 minutes in 4% formaldehyde in PBS, washes 3 x 10 minutes in PBT, stains with DAPI 5 minutes, washes 3 x 10 minutes in PBT and coverslipped.  Macroscopic evaluation of the section shows good architecture, but microscopically the cell morphology isn't good, especially the retina.  The eye globes are received without the cornea, so I filled the anterior chamber of the eye with OCT, then surrounded the remainder of the globe with OCT in a Peel-away mold.  I then froze it in liquid nitrogen-cooled isopentane in an attempt at minimizing freezing artifact.  Any suggestions on improving our cellular morphology would be greatly appreciated.
Teri Johnson
Managing Director Histology Facility
Stowers Institute for Medical Research
1000 E. 50th St.
Kansas City, Missouri  64110

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