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>Has anyone heard of FACS-gal, FDG, Fluoreporter substrate used for lac-z
>localized to the nucleus in live cells? Is this even possible using
>hypotonic treatment?
>
>Just doing a reality check for a flow user.
>
>Thanks,
>Rochelle Diamond
>Caltech Biology
>Flow Cytometry/Cell Sorting Facility
>
I replied
>We stained cells successfully with FDG that had nuclear localized lac-z.
>There were no problems with the standard assay, and using a fixation like
>Clevenger's (low formaldehyde/triton X-100), we had better retention than
cells >expressing normal lac-z.
This message was hurried and not well thought out. We were able to detect
nuclear localized lac-z with FDG staining of live cells by the published
methods. When we fixed cells with Clevenger's procedure, we had better
retention of lac-z, as detected by monoclonal antibody staining of lac-z,
compared to cells expressing normal lac-z. The previous message implies
that the substrate was retained within the cells and that is not likely to
be the case. We have not tried to find out. Sorry for any inconvenience.
Jake
James (Jake) Jacobberger
Associate Professor
216-368-4645 phone
216-368-3432 fax
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