[Histonet] (no subject)

[jason m]

Questions about LCM.

I am attempting a unique application of Laser Capture. In order isolate pure 
ovarian surface epithelium I am taking the ovary directly in the OR and 
rolling it between two glass slides. The surface epithelium easily detaches 
and sticks (loosely) to the slide. I then follow this protocol for staining 
and dehydration.

slides are frozen in the OR.
5-10 minutes acetone/EtOH fixation (3:1)
30s dH2O
1s hematoxylin
30s dH2O
graded EtOH 1 minute each
5 minutes xylenes
airdray
proceed with LCM

The probem I am having is that the cells sometimes become too firmly 
attached to the slide and will not under any means come off the slide with 
the LCM scope. I do not have a lot of material to play with so I would 
appreciate any help anyone might have. Will a longer xylenes incubation 
detach the cells or a shorter fixation? I need the cells attached firmly 
enough that they dont come off in the solutions but not so firmly attached 
that they wont come off the slide via LCM. Does anyone have experience with 
this type of thing?

Thanks for any help.

Jason.

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