C. J. Buckley

Illumination for coherent soft X-ray applications: the new X1A beamline at the NSLS.

The X1A soft X-ray undulator beamline at the NSLS has been rebuilt to serve two microscopy stations operating simultaneously. Separate spherical-grating monochromators provide the resolving power required for XANES spectroscopy at the C, N and O absorption edges. The exit slits are fixed and define the coherent source for the experiments. The optical design and the operational performance are described.

Examination of the penetration of polymeric methylene di-phenyl-di-isocyanate (pMDI) into wood structure using chemical-state x-ray microscopy

Summary The penetration behaviour of isocyanate-based wood resins was evaluated using x-ray microscopy. Aspen wood pieces were bonded together in a controlled manner. These were embedded in a methacrylate-based resinand thin sections were prepared, cut from the transverse face of thewoodcomposite. X-ray images of these sections were prepared at several selected x-ray energies to allow the isocyanate, cellulose, lignin and the embedding agent distributions to be mapped. The isocyanate resin was found to penetrate deeply into the wood. The resin enters large cell lumen, and wicks along the inner cell wall surfaces. The resin accesses connected cells via connecting pits, which become filled with the resin. The affinity of the isocyanate to the inner surfaces of the large cells is probably due to the hydrophobicity of these surfaces. Isocyanate resins do not penetrate into the smaller parenchyma and tracheid cells and indeed do not even wet the inner surfaces of these cells where isocyanate entry has been allowed due to damage of the cell at the macroscopic surface of the wood. If isocyanates penetrate into the wood-cell walls of the large cells, the concentration in the cell walls has been determined to be less than 2%of the bulk concentration. This lower limit is the sensitivity limit imposed by photon statistics in the data.

The effect of soft X‐radiation on myofibrils

Myofibrils, the contractile organelles from striated muscles, have been examined in the X‐ray microscope to determine the effect of radiation on their function and structure. Using X‐rays of energy 350–385 eV in the water window we find that after an exposure to 7·5 × 105 photons/μm2 (calculated to give an absorbed dose of 20000 Gy) the myofibrils will no longer contract. The use of the free radical scavenging agent, DMSO, gives some protection to the fibrils. It has also been found that after this much irradiation the fibrils lose up to 20% of their mass. Further substantial mass loss occurs on subsequent irradiation. After 25 times the loss‐of‐function exposure only 30% of the mass remains. Analysis of a series of images of the same myofibril covering this range of exposures shows that the mass is preferentially lost in some areas of the structure and consequently significant structural changes occur.

THE SCANNING TRANSMISSION MICROSCOPE AT THE NSLS

Abstract The scanning transmission soft X-ray microscope (STXM) that has been under development at the National Synchrotron Light Source [H. Rarback et al., Rev. Sci. Instr. 59 (1988) 52] has been substantially ungraded for operation with the X1 undulator [C. Buckley et al., Rev. Sci. Instr. 60 (1989) 2444]. The principal new features are: optical prefocusing, using a visible light interferometer; a dedicated VAXstation 3200 with a more user friendly and flexible software system for image acquisition and analysis; a flow cell that makes it possible not only to keep the specimen wet during exposure, but to change the fluid around the specimen as well; and a more compact proportional counter that is capable of counting rates of several MHz. In conjunction with new zone plates of better resolution and higher efficiency [E.H. Anderson, SPIE 1160 (1989) 2], the microscope is ready for a period of extended use in biological imaging.

X1A: Second-generation undulator beamlines serving soft x-ray spectromicroscopy experiments at the NSLS

The X1A undulator beamline is being rebuilt with two separate monochromators on its two branches. The new arrangement will deliver spatially coherent beams to imaging experiments, with spectral resolving power of up to 5000, and the capability to optimize the resolving power versus flux. The beamlines will operate simultaneously, and each will use 15 percent of the undulator beam, yet deliver high coherent flux. Because of the small beam divergence, the spherical grating monochromators can operate with fixed exit arms throughout the 250–800 eV range.

Coherent radiation for x-ray imaging—The soft x-ray undulator and the X1A beamline at the NSLS

An undulator-based beamline was built and commissioned at the National Synchrotron Light Source to provide tunable coherent radiation in the 200-800 eV range. The low emittance of the storage ring means that the undulator source has high brightness so that a large flux of coherent x rays is delivered to experimental stations. The beamline uses a horizontally dispersing bichromator that allows two experiments to run simultaneously, making use of the first and second harmonics of the undulator output. In addition, the use of horizontally deflecting optics enables the beamline alignment to be insensitive to electron beam motion since the horizontal electron beam size is quite large. The beamline and its performance are discussed with emphasis on the optics and on stability, radiation, and vacuum considerations.

Soft X-Ray Absorption Imaging of Whole Wet Tissue Culture Cells

Using the Stony Brook/NSLS Scanning Transmission X-Ray Microscope (STXM), we have produced high resolution images of whole, wet, fixed and unfixed fibroblasts. Images of unfixed cells were taken at a wavelength of λ=34.1A. An image can take 5 minutes to achieve a 14:1 background noise. The x-ray dosage, required to make such an image on wet initially live fibroblasts does not cause immediate observable structural damage to the target cells.

THE MEASURING AND MAPPING OF CALCIUM IN MINERALIZED TISSUES BY ABSORPTION DIFFERENCE IMAGING

This paper reviews the mapping of calcium in mineralized tissues by absorption difference imaging using scanning x‐ray microscopy at the calcium L edge. In particular, the mapping of embedded and sectioned mineralized tissues is considered. Here, the technique is discussed in detail and the accuracy is evaluated with and without corrections. The technique is considered under two scenarios: the use of differences in absorption cross section at pre‐ and postionization energies, and the use of differences arising from near‐edge x‐ray‐absorption features.

Soft X-ray Microscopy in Biology and Medicine: Status and Prospects

There are two central motivations for developing new scientific methods. One is, of couse, to accomplish what established methods cannot. A second is for comparison: To verify the conclusions of established methods. That is, are results obtained by one method congruent with those obtained by another independent means of measurement? In regard to microscopic imaging in biology, this means that we seek to ground our view of microscopic structure on more than a single methodological standard, with whatever particular uncertainties that standard presents. These are the motivations that underlie the current impetus for the development of x-ray microimaging methods. Our knowledge of the internal structures of biological cells has been shaped in great part by 40 years of study applying and developing the methods of electron microscopy. This has led to the evolution of a model of the cell that contains defined structures with established details and known spatial relationships. Belief in the fidelity of this model to the natural cell rests in great part on the understanding that the preparative procedures commonly used in electron microscopy, procedures that greatly modify the natural object, do not alter or distort intracellular structure as to form, location or high resolution detail. Even though the cell as seen in the electron microscope most certainly resembles the natural object, important questions of the faithfulness of the image often remain.

X-ray Microscopy with the NSLS Soft X-ray Undulator

Soft x-rays are attractive for microscopy because they tend to be less damaging to specimens than charged probes. In addition their interactions with specimens can result in element-specific information and the penetration depth is adjustable by choice of the beam energy. Resolution on the order of 50 nm has been demonstrated, and further improvements are anticipated. The experimental program at the NSLS X1A beamline is dedicated to soft x-ray microscopy. We are developing two types of instruments, both dependent on the remarkable brightness of the undulator source. One of these uses a Fresnel zone plate to focus the beam to a small size. This microprobe is used either to study biological specimens, or, in a different apparatus, to study surfaces by photoelectron microscopy. The other type of instrument makes use of x-rays diffracted by the specimen. In this category we are involved in the development of Gabor holography, Fourier transform holography, and in imaging by soft x-ray diffraction.