Automated outlier rejection in scaling will handle a lot of things,
including ice. Works better with high multiplicity. Unless, of course,
your ice rings are "even", then any integration error due to ice will be
the same for all the symmetry mates and the scaling program will be none
the wiser. That said, the integration programs these days tend to have
pretty sensible defaults for rejecting spots that have "weird"
backgrounds. Plenty of structures get solved from data that has
horrible-looking ice rings using just the defaults. In fact, I am
personally unconvinced that ice rings are a significant problem in and
of themselves. More often, they are simply an indication that something
else is wrong, like the crystal warmed up at some point.
Nevertheless, if you suspect your ice rings are causing a problem,
you can try to do something about them. The "deice" program already
mentioned sounds cool, but if you just want to try something quick,
excluding the resolution ranges of your ice rings can be done in sftools
like this:
select resol > 3.89
select resol < 3.93
absent col F SIGF DANO SIGDANO if col F > 0
and repeat this for each resolution range you want to exclude. Best to
get these ranges from your integration program's graphics display.
In mosflm, you can put "EXCLUDE ICE" on either the "AUTOINDEX" or
"RESOLUTION" keywords and have any spots on the canonical hexagonal ice
spacings removed automatically. The problem with excluding resolution
ranges, of course, is that your particular "ice rings" may not be where
they are supposed to be. Either due to something physical, like the
cooling rate, or something artificial, like an error in the camera
parameters. It is also possible that what you think are "ice rings" are
actually "salt rings". Some salts will precipitate out upon
cryo-cooling. Large ice/salt crystals can also produce a lot of
non-Bragg scatter, which means that you can get sharp features far away
from the resolution range you expect. On the other hand, if you have
cubic ice instead of hexagonal ice (very common in MX samples), then
there are no rings at 3.91A, 3.45A, 2.68A and throwing out these
resolution ranges would be a waste.
Another way to exclude ice is to crank up background-based rejection
criteria. In denzo/HKL2K, you do this with the "reject fraction"
keyword, and in mosflm, REJECT MINBG does pretty much the same thing.
There are lots of rejection options in integration programs, and which
one works in your particular case depends on what your ice rings look
like. Noone has written a machine-vision type program that can
recognize and handle all the cases. You will need to play with these
options until the spots you "don't like" turn red in the display.
Of course, the best way to deal with ice rings would be to inspect each
and every one of the spots you have near ice rings and decide on its
intensity manually. Then edit the hkl file.
Which brings me to perhaps a more important point: What, exactly, is the
"problem" you are having that makes you think the ice rings are to
blame? Can't get an MR solution? Can't get MAD/SAD phases?
Ice has a bad rep in MX, and an undeserved one IMHO. In fact, by
controlling either cryoprotectant concentration or cooling rate
carefully, you can achieve a mixture of amorphous and cubic ice, and
this mixture has a specific volume (density) intermediate between the
two. Many crystals diffract much better when you are able to match the
specific volume of the stuff in the solvent channels to the specific
volume protein lattice is "trying" to achieve on its own. A great deal
of effort has gone into characterizing this phenomenon (authors: Juers,
Weik, Warkentin, Thorne and many others), but I often meet frustrated
cryo-screeners who seem to have never heard of any of it!
In general, the automated "outlier rejection" protocols employed by
modern software have taken care of most of the problems ice rings
introduce. For example, difference Pattersons are VERY sensitive to
outliers, and all it takes is one bad spot to give you huge ripples that
swamp all you peaks, but every heavy-atom finding program I am aware of
calculates Pattersons only after fist doing an "outlier rejection"
step. You might also think that ice rings would mess up your preciously
subtle anomalous differences, but again, outlier rejection to the rescue.
Now, that said, depending on automated outlier rejection to save you is
of course a questionable policy, but it is an equally bad idea to
pretend that it doesn't exist either. It is funny how in MX we are all
ready to grab our torch and pitchfork if we hear of someone manually
editing their hkl files to get rid of reflections they "don't like", but
as long as "the software" does it, it is okay. Plausible deniability
runs deep.
-James Holton
MAD Scientist
On 10/11/2011 8:16 AM, Francis E Reyes wrote:
All,
So I have two intense ice rings where there appear to be lattice spots in
between them.
I understand that any reflections that lie directly on the ice ring are
useless, however, how do software programs (HKL2000, d*Trek, mosflm, XDS) deal
with these intermediate spots?
It would seem to me that employing a 'resolution cut off' just before the ice
ring (on the low resolution side) would be improper, as there are spots on the
high resolution side of the ice. (see enclosed .tiff)
In fact, how do these programs deal with spots lying on ice rings? Are they
rejected by some algorithm by those programs during integration, or is it up to
the scaling/merging (by SCALA for example) step to deal with them?
Thanks!
F
---------------------------------------------
Francis E. Reyes M.Sc.
215 UCB
University of Colorado at Boulder
