discussion on strategies introductory notes - omega vs. phi scans - beam polarization - single sweep...
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Discussion on StrategiesIntroductory Notes
- omega vs. phi scans
- beam polarization
- single sweep vs. multi sweep
- xtal shape as re-orientation/re-centering factor
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ˆ J ( h ) 1
s J u (h )Exp( h B h T )
Reconstruction of the mean reflection intensities using limited experimental data set:
<E2> profiles – a feature of PROTEINS, NOT APPLICABLE TO SMALL MOLECULES
10 5 3.33 2.5 2 1.67 1.43 1.25 1.11 1 0.9099
9.5
10
10.5
11
11.5
12
12.5
resolution d (A)
ln <I>
(a)o
1hq3 : [ ]=0.63, []=0.06
1at0 : [ ]=0.00, []=0.60
1d5t : [ ]=0.27, []=0.23
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Optimization target: Signal/Noise
• NOT the time to be spent for experiment, number of frames to collect, etc …
• ALL the data collection parameters (multi- sub-wedge, variable exposure time, etc.) are optimized simultaneously -
Example: multiplicity vs exposure time
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Radiation Damage -
• Compensation of intensity decay by adjusting (increasing) the exposure time / frame is essential :
Total dose per data set is not important– defined by the long exposure of the LAST
frames– short exposures of the FIRST frames are
critical
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What works in BEST now?optimal orientation with respect to:
• Overlaps (~90% of failing experiments – J. Holton )
- also with isometric cells @ high mosaicity
• Intrinsic diffraction anisotropyeach diffraction pattern is maximally isotropic,
S/N in a weak direction compensated by exposure (small effect when judged by standard "resolution shell" statistics)
• Low noise in anomalous difference dataanomalous difference error model (radiation inducednon-isomorphism) accounts for the difference in dose between the observed Bijvoet mates
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Minimal RFriedel= <|<E+>-<E->|> vs. Resolution and Orientation (error
contribution to the difference only, no anomalous scattering contribution
P2
0
2
4
6
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16
0 0.05 0.1 0.15 0.2 0.25
1/d^2
Rfr
ied
el, %
0 1 01 0 01 1 0Random
P222
0
2
4
6
8
10
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16
0 0.05 0.1 0.15 0.2 0.25
1/d^2
Rfr
ied
el,
%
random1 0 01 1 01 1 1
P4
0
2
4
6
8
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16
0 0.05 0.1 0.15 0.2 0.25
1/d^2
Rfr
ied
el, %
0 0 11 0 0 1 1 1
P23
0
2
4
6
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0 0.05 0.1 0.15 0.2 0.251/d^2
Rfr
iede
l, % 1 0 0
1 1 01 1 1random
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Data collection using multiple crystals
Reference images
Auto-indexing
BESTCrystal characterization and ranking
Determination of maximal achievable resolution
Optimal crystal orientation(s)
Experimental aim
Crystal 3Crystal 5
Crystal 1Crystal 8
D.C. planCompleteness 23%
Completeness 58%
Completeness 91%
Completeness 99.7%
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Omega vs. Phi scans
Omega scans - orientation wrt scan axis is optimized
OverlapsRadiation-induced non-isomorphismMulti-crystalsAAS
Phi scans- orientation wrt BEAM (direction/electric field vector) is varied
"true redundancy" (– no advantage wrt. Omega,but - may be - less limitations)
Blind region reduction ( - when in a symmetric setting) AAS?
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Beam polarization• Isotropic scattering –
Scan axis || Electic Filed vector is optimal, though only important at high resolution ( < 2*wavelength)
Vertical OMEGA is of advantage for the microbeam (gravity)
PHI is mechanically non-micro
• AAS
BEST minimizes the noise in anomalous diffrence data (fully applicable to AAS data)
the target describing the AAS signal is required
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Single Sweep vs. Multi Sweep
Multi sweep on a single crystal:
Blind region completionMultiplicity
Partial data set completion (disaster scenario)
From the point of view of implementation in BEST, Multi-Sweep strategy is a particular case of multiple crystal data collection optimization with the goniometric limitations
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Single Sweep vs. Multi Sweep• "Fast" coverage of an asymmetric unit on a
single crystal – no advantage in signal-to-noise!
Single sweepRadiation damage
Disadvantage – Inhomogeneous S/N
Single sweepRD compensation
Multiple sweeps
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xtal shape as re-orientation/re-centering factor
• Exploiting ALL of the crystal volume is critically important
• Severe mismatch of Xtal/Beam size – major limitation to sample characterization,
strategy and data quality in general
• Use Kappa to match the Xtal/Beam size
(at least in a vertical direction), Simplify line scans along Omega