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It is my understanding that the "ITC" in FITC stands for
isothiocyanate, which will spontaneously bind the fluorescein to free
amino groups -such as the termini of proteins.   I don't know what
other amino compounds might be present in the dough, though.
Regardless, I would have expected to rinse out the excess FITC unless
the initial amount was quite low.

Joel


> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Dear all,
>
> Apologies in advance for double post to confocal and microscopy list.
>
> The group studying flour and dough structure here wants to use FITC to
> stain the dough protein so they can get an idea of the macrostructure
> of the dough.  The protocol has been published in a couple of papers
> from another group, and after staining the blob of dough they looked
> at it under confocal to see changes in fluorescence pattern after
> various amounts of mixing, and assumed that any fluorescence came from
> protein aggregates.  This worries me, as I wasn't aware that FITC per
> se was such a specific stain for protein, and the published protocol
> says nothing about washing out excess free dye.  The alternative I
> thought of was to use fluorescamine, which is only fluorescent when
> bound to protein, so you could put your sample into the dye and
> observe without rinsing.  (Needs UV, though.)
>
> Does anyone have experience with this sort of material, comments about
> FITC as a protein stain?
>
> TIA,
> Rosemray
>
> Dr. Rosemary White              [log in to unmask]
> Microscopy Centre                 ph.     61-2-6246 5475
> CSIRO Plant Industry            mob.  61-0402 835 973
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> Australia


Joel B. Sheffield, Ph.D.
Biology Department, Temple University
1900 North 12th Street
Philadelphia, PA 19122
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(215) 204 8839, fax (215) 204 0486
http://astro.temple.edu/~jbs