Microcrystallography using single-bounce monocapillary optics

Abstract

X-ray microbeams have become increasingly valuable in protein crystallography. A number of synchrotron beamlines worldwide have adapted to handling smaller and more challenging samples by providing a combination of high-precision sample-positioning hardware, special visible-light optics for sample visualization, and small-diameter X-ray beams with low background scatter. Most commonly, X-ray microbeams with diameters ranging from 50 microm to 1 microm are produced by Kirkpatrick and Baez mirrors in combination with defining apertures and scatter guards. A simple alternative based on single-bounce glass monocapillary X-ray optics is presented. The basic capillary design considerations are discussed and a practical and robust implementation that capitalizes on existing beamline hardware is presented. A design for mounting the capillary is presented which eliminates parasitic scattering and reduces deformations of the optic to a degree suitable for use on next-generation X-ray sources. Comparison of diffraction data statistics for microcrystals using microbeam and conventional aperture-collimated beam shows that capillary-focused beam can deliver significant improvement. Statistics also confirm that the annular beam profile produced by the capillary optic does not impact data quality in an observable way. Examples are given of new structures recently solved using this technology. Single-bounce monocapillary optics can offer an attractive alternative for retrofitting existing beamlines for microcrystallography.

Figure 1

Cross-sectional view of a capillary segment with elliptical internal profile. X-rays from a divergent point source located at the left focus of the ellipse converge to the opposite (right) focus after reflecting at a glancing angle from the internal surface. The maximum divergence of the focused beam, θ_div, results from rays collected from opposite sides of the tube. A second smaller divergence, θ_m, is from the elliptical curvature itself. Focal length F, capillary length L _c and distance to source L, together with maximum divergence, fully specify the capillary design. Parameter r measures the radial distance from the center of the focal spot within the focal plane (dotted line). The scale is highly compressed for the purposes of illustration since L >> F.

Figure 2

Distortion of the capillary figure owing to gravity. Calculations from the theory of flexure of simple beams (Appendix A ) show that proper mounting is an important factor if capillary figure errors are to be reduced to a level appropriate for next-generation X-ray sources. The y-axis represents deviations from the ideal centerline of the optic. Support points are shown as triangles, with open triangles corresponding to the dashed line. The solid line supported at the solid triangles represents an optimally supported optic which minimizes overall figure error.

Figure 3

Design and mounting of capillary optics. A precision-bored barrel (a) encloses the glass capillary, which is supported by two location discs of precisely defined aperture. Gentle pressure is applied using a plug with viton cushions to hold the optic in place. A guard aperture to remove parasitic scatter is positioned at the end of the enclosure about 10 mm from the tip of the capillary. The barrel is mounted on a movable base (b) controlled with a piezomotor (red in the lower left) from New Focus, San Jose. A black registration mark on the end of the barrel (right) marks the tip of the capillary and is used for manual positioning. The sample is mounted on the yellow plastic base at the far right.

Figure 4

Diffraction spot profile as a function of detector distance. Diffraction spots close to the beamstop begin to show the characteristic annular microbeam profile once the sample-to-detector distance exceeds about 500 mm. This (−2−21) reflection, which occurs at 18.8 Å, was produced by a large 100 µm-diameter good-quality lysozyme crystal. The upper row of images has been contrast-enhanced to make the halo visible. The lower row of surface plots has been scaled to uniform height for easy comparison. In actuality, the 136 mm profile is six times taller than the 876 mm profile.

Figure 5

Far-field beam profile image and visualization camera. At large distances (>500 mm) from the capillary tip, the X-ray microbeam is annular in form as seen by fluorescence on a CdWO₄ crystal (a). The direct unfocused beam is the disc at the center while the focused beam has diverged into a halo. Black grid lines mark 1 mm squares. A mini video camera equipped with optical spacers and CdWO₄ crystal is fixed to the protective ‘garage door’ shield on the crystallography CCD detector and can be easily moved into place to check the alignment of the X-ray optic (b).

Figure 6

Terbium-doped glass fiber for aligning the sample spindle with the X-ray microbeam. X-ray-sensitive scintillating glass is easily drawn into fibers of diameter 30 µm or smaller. A cross-hair is also shown in the figure with tick marks every 10 µm. The broad green glow has a 40 µm base corresponding to the exit diameter of the capillary tip, and the focused radiation is visible as a bright spot at the center.