Hello everybody,
I am dealing with the standard problem that I have (spherical) crystals and I
want to completely make sure that it is nothing else than protein. Since the
amount of protein I am working with is very limitated, I
want to get the maximum of information out of the few xtals I have.
So we put them into the beam (Rigaku, Saturn 944 CCD, 1min exposure time) and
there was no single spot (no salt spots either). Due the stability of the
xtals, I am not so sure about soft/hard (poking with needle), IZIT did not show
much, either. I did the same crystallization experiment, again but without
protein (vapour/diffusion: liquid drops so far). Another method is to try to
wash the xtals and do an SDS-PAGE afterwards, but this destroys the crystals
(maybe already during the washing process in the worst case). Before I do this
procedure, I wonder if it would make sense to try to dye them via another
method than IZIT. However, glutaraldehyde should mess up the SDS-PAGE plan
because of the crosslinking effect (right? no "unlinking" possible?).
Since I do not have any practical experience with these two techniques, I
wonder what priority you would suggest, SDS-PAGE or glutaraldehyde? Or is there
another method (like unsuccessful staining with IZIT) I am not aware of, that
would allow an SDS-PAGE or staining by glutaraldehyde afterwards? Many thanks
for your suggestions!
Jan
