Department X-Ray Microscopy
Technique Introduction
Shown is an example workflow for use of X-ray tomography.
The transmission X-ray microscope is a bright-field microscope that operates at a wavelength of 2.4 nm. This wavelength enables super-resolution bright-field images, and it also provides for a natural contrast without specimen staining because photons at this wavelength are strongly absorbed by organic matter compared to water.
The objective lens known as a zone plate is constructed by nanofabrication. It is composed of a series of concentric rings of nickel or gold at a pre-calculated spacing designed to diffract the X-ray light to a set of focal points known as the diffraction orders. A typical zone plate diameter is only a few hundred microns. The width of the outermost zone, typically in the range of 15-50 nm, determines the resolution of the zone plate objective.
Typical zone plate objective lenses for X-ray microscopy have a large depth of focus spanning at least several microns, even though the xy resolution is on the order of nanometers. As a result, the image of a typical tissue culture cell is a projection image.
3D images of a specimen can therefore be obtained by tilting the specimen through a series of angles and then performing a tomographic reconstruction.
Synchrotrons are typically used to generate sufficiently bright illumination for an X-ray microscope, although table top microscopes are also available using laser plasma sources.
Shown is an axial (z) and radial (r) profile of a typical X-ray microscope PSF (40 nm zone plate objective). Not drawn to scale as the radial distance is greatly magnified compared to the axial distance.