Neurolucida and Fluoview FV10i - (or general imaging help)

Does anyone have experience imaging clarified tissues with either of these machines? If not, maybe you can help me anyway?

I'm having significant issues getting clear and focused images. The brain section I'm using is ~500μm, and Hoechst stained. It was passively cleared in a water bath (37°C) for about 6-8 weeks.

There was endogenous GFP (d2GFP), but that seems to have degraded or washed out. My PI is concerned that the cells degraded after 2-3 months of waiting time. I think my issue lies in mounting and with the FocusClear. I've been using a slide and cover slip (with bluetack), and the results have been less than impressive.

Any ideas? Thanks a lot!!


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    LinusMGLinusMG Posts: 20
    My guess is that probably the problem will come from the objective you use and the refractive index mismatch between objective and sample. What are the depths of the images?
    Do you have any pictures of the slide and coverslip with the sample? (does it look transparent?)
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