staining a chargeless protein

[Phillip J Robinson]

Olin Anderson <oandersn at pw.usda.gov> wrote:

>We are working with a 55 kDa polypeptide that has no charged amino acids.
>Attempts to stain the polypeptide on PAGE gels has failed - tries
>included commasie blue, amido black, silver, copper stain, Sypro, 
>anilionaphthalene sulfonate, and fluorescein isothiocyanate.  The only
>amino group is the N-terminal and it seems unreactive/blocked.  The
>only amino acids in the polypeptide are P, G, Q, Y, T, and S.  Any
>suggestions how to stain/detect this polypeptide in a gel?

It seems simple enough to me.  Here's two ideas for negative staining.
However, you seem to have tried them already.  I have to say that if
both fail I also feel that you may have not got any "penetration" into
the gel of your protein or one of the following really should work
(shouldn't they?).  Both the following do NOT depend on the charge of
the protein to the best of my understanding, just on the fact that a
protein occupies space in the gel.

Try copper chloride (sulphate?) staining.  It apparently stains the
background, by turning it opaque, and leaves the protein
clear/unstained.  View on a black sheet of paper.  I've tried it,
seems more sensitive than coomassie, as originally claimed.  I don't
have the reference handy, but will email upon request.

Alternatively, we have often observed that if one overstains a
silver-stained gel, some proteins that were not stained show up
beautifully clear against the overstained background.  

Anyone else out there with other ideas on such negative staining
approaches?

PS:  I too am curious about the "natural" occurence of such a protein.
Care to elaborate?

Phil,				Newcastle, Australia