[Histonet] Problems with rhodanine

[Katherine Cano]

Hi all,

I'm a graduate student in a neuropsychology lab, and we are having some
problems with our histology. I was hoping someone might have some advice.

My lab is doing staining for metals in fresh frozen mouse brain, sectioned
at 10u or 20u. Some of our animals are fed high copper supplements and we
see high copper in areas of our tissue with other methods, so we are trying
to pilot rhodanine for copper. However, the protocols we have for rhodanine
give conflicting instructions.

We have unfixed, fresh frozen tissue, which is part of the problem, since
ALL our protocols are for FFPE tissue. Two of the protocols call for
microwave method, but we don't have an appropriate microwave so we are using
the conventional oven method, which says to heat the slides for 18 hours at
37 or 30 degrees centigrade.

When we looked at the coverslipped slides, it looked as though the tissue
had washed off in place. I have seen this happen before with immuno, and my
histology professor said it was a problem with the slide subbing; however,
we are using Erie Superfrost Plus slides, which I did not think would have
that problem. We DID end up moving the slides between buildings, so maybe
the tissue was jostled too much during transport and that's why it washed
off?

Does anyone have any experience with rhodanine on fresh frozen tissue? Do
you need a special protocol?
 Does it just not work? Any input would be appreciated.

Thanks in advance,
Katherine