Thanks, Ron,
Regarding the "bathing" question, these days the major source of error we
find in synchrotron-based data is crystal damage. Several groups, notably
the two ID23s (one each of pairs of matched canted undulators at ESRF and
APS) are producing small x-ray beams, on the order of 5-10 micron
diameters, and are saying that the principal use to which they're put is
to shoot a single larger crystal several times. There's a certain synergy
here -- each new exposure is essentially a new and undamaged tiny crystal,
and the orientations of each succesive spot on the crystal are essentially
identical. The orientations abut nearly precisely and various
data-reduction questions are simplified. These days, where x-tals tend to
be smallish and synchrotron access is pretty available, this seems to us
to be an excellent strategy.
Regarding absorption corrections, here at the PXRR/NSLS a whole lot of
data are taken with wavelengths on the order of one Angstroem. At these
wavelengths the absorption of a typical crystal in its mount, even one
that is, say 3M ammonium sulfate, is nearly negligible. On the other
hand, if one tries to optimize S or I anomalous with 1.7 Angstroem
radiation, all bets are off. Here I'm sure >you< would take excellent
data.
About empirical corrections: to employ Tony North's method in about 1969 I
took what are formally psi scans (repeat mesurements on a diffractometer
of one reflection at increments of rotation about one real-lattice vector)
on the trypsin/soybean-trypsin-inhibitor complex. I was using "Wyckoff"
scans* to do this. The crystal was an orthorhombic needle, and I was
rotating about the needle axis. I observed that the rocking curve varied
from narrow to wide, with the extremes being separated by about 180 deg.
I'd jammed the needle into a tapered capillary and it was slightly
bent(!), giving a focused beam in one direction (sharp) and an unfocused
one in the opposite (broad). I'm told this happens these days with frozen
needles sticking out of the gob of vitreous mother liquor.
* Hal Wyckoff (the inventor of the true Inverse Beam Method**) reasoned
that there were three ways to "integrate" a reflection: one could rotate
the crystal through its rocking curve (the rotation method), one could use
a broad bandpass of wavelengths (the Laue method), or one could use a
widely convergent source (the Wyckoff scan). To do this one increased the
take-off angle of the x-ray tube (Ron knows what this is, but those of you
who don't know should ask the oldest crystallographer nearby) so that the
angle subtended from the crystal would fill the rocking curve. In
principal the single largest measurement would equal the integrated
intensity. To hedge ones bets, one did a five-point scan and summed the
top three. This was how I could see the sharpness of the scans.
** Wyckoff also invented a diffractometer that had two opposing x-ray
generators, literally shooting at the crystal from opposite directions,
and then two opposing detectors, each on its own 2-theta arm. One
detector would measure (h,k,l) at precisely the same time the other would
measure (-h,-k,-l). What we do now, to flip the rotation axis by 180
degrees, seeing the (-h,-k,-l) reflections in a mirror image to the
(h,k,l) ones, would better be termed the Friedel Flip.
Keep those cards and letters coming, folks.
Bob
On Sun, 25 Nov 2007, Ronald E Stenkamp wrote:
Just a few comments on "consider a crystal bathed in a uniform
beam".
I've not fully bought into the idea that it's OK to have the
beam smaller than the crystal. I learned most of my crystallography
in a lab dedicated to precise structure determinations, and somewhere
along the line, I picked up the idea that it's better to remove systematic
errors experimentally than to correct for them computationally.
(Maybe that had something to do with the computing power and programs
available at the time?)
Anyway, I thought the reason people went to smaller beams was that
it made it possible to resolve the spots on the film or detector. Isn't that
the main reason for using small beams?
In practice, I guess the change in crystal volume actually diffracting
hasn't been a big issue and that frame-to-frame scaling deals with
the problem adequately.
I'm less convinced that frame-to-frame scaling can correct for absorption
very well. Due to our irregular-shaped protein crystals,
before the area detectors came along, we'd use an empirical correction
(one due to North comes to mind) based on rotation about the phi axis
of a four-circle goniostat. It was clearly an approximation to the
more detailed calculations of path-lengths available for crystals with
well-defined faces not surrounded by drops of mother liquor and glass
capillaries. Has anyone checked to see how frame-to-frame scaling
matches up with analytical determinations of absorption corrections?
It'd be interesting to determine the validity of the assumption that
absorption is simply a function of frame number.
Ron
--
=========================================================================
Robert M. Sweet E-Dress: [EMAIL PROTECTED]
Group Leader, PXRR: Macromolecular ^ (that's L
Crystallography Research Resource at NSLS not 1)
http://px.nsls.bnl.gov/
Biology Dept
Brookhaven Nat'l Lab. Phones:
Upton, NY 11973 631 344 3401 (Office)
U.S.A. 631 344 2741 (Facsimile)
=========================================================================
