High resolution X-ray transmission radiography
The common methods of structure imaging of small animals include optical microscopy, electron microscopy and fluorescence microscopy. These techniques provide detailed information about external and internal anatomy. However, the sample preparation is usually laborious, time consuming and often also expensive. Moreover, the samples, after being examined with one of the mentioned procedures, cannot be used for further purposes. Therefore, new approaches are needed and are being developed. Among them high resolution X-ray radiography, which is often exploited also in tomographic mode to provide 3D images, appears as a promising technique for real time and nondestructive visualization. This method enables as well in-vivo analysis of time dependent physiological processes in living specimen.
X-ray transmission radiography is based on beam intensity attenuation. A fraction of the beam has to be absorbed by the sample in order to obtain an image of its structure. The imaging of biological samples consisting of soft tissue is particularly difficult due to low differences in X-ray attenuation. New X-ray imaging detectors like Medipix2 with unlimited dynamic range allow to reach good contrast and, in combination with advanced point like X-ray sources, it is possible to refine spatial resolution of the method below 1m.
This contribution describes an arrangement for X-ray transmission radiography with the state-of-the-art semiconductor hybrid pixel detector Medipix2 (512 x 512 pixels with 55μm pitch) and a commercial X-ray point source. The main advantages of the Medipix2 pixel detector include: high sensitivity to low energy X-ray photons; position sensitive and noiseless single photon detection with preselected photon energies; photon counting in each pixel performed by digital counter, therefore with no dark current; digital integration providing unlimited dynamic range and absolute linearity in device response to number of photons; high speed digital communication and data transfer.

X-ray phase sensitive imaging
Conventional transmission radiography is based on beam intensity attenuation. A fraction of the beam has to be absorbed by the sample in order to obtain an image of its structure. The sensitivity of this method is limited for weakly absorbing objects such as biological samples consisting of soft tissue. Phase sensitive imaging on the other hand uses phase shift of the photons passing the sample. Variations in the phase result from changes in the refractive index across the medium. A substantial advantage of this method is that X-ray photons used for phase shift imaging penetrate through a sample and therefore they do not contribute to radiation dose.
There are several approaches of the phase shift visualization. An interferometric approach uses interference of the transmitted beam with the primary beam. A diffractometric method (DEI) uses an analyzer crystal to separate transmission and phase image. And a so called in-line method separates these beams at a large object to detector distance.
Although phase sensitive X-ray imaging presents many advantages it is not routinely used in biological research. Realization of the method currently demands high intensity and highly coherent X-ray beams which are accessible mainly at large scale and costly synchrotron facilities. In addition the use of perfect crystals is usually required both to produce such highly coherent beams and to analyze the transmitted images. Avoiding the need for such complex instruments and large scale facilities, it has been shown that in-line phase enhanced imaging can be carried out also with polychromatic microfocus X-ray tubes. The main problem of such systems is the low beam intensity which prolongs exposure time to such an extent that common digital imaging detectors (CCD, Flat panels) are disadvantageous due to their low efficiency, dark current and noise.
This contribution presents a compact table-top phase contrast enhanced imaging system based on a microfocus X-ray tube and the position sensitive single photon counting pixel detector Medipix2. Each pixel of the detector is provided with preamplifier, pair of discriminators and counter. Discriminators allow full suppression of the noise and selection of energy range of interest. Combined with its high detection efficiency, this feature makes the device ideal for in-line phase enhanced imaging. The spectral sensitivity of the detector together with the polychromatic nature of the beam allows to distinguish an attenuation image from a phase (refractive) image. Spatial resolution of the system can be on the submicrometer level, measuring times less than a minute and doses of order of mGy.
Applications of the system such as the investigation of internal structure of several biological samples (termites, mouse kidneys and lungs) including 3D reconstructions will be presented. The simplicity of the system allows for routine laboratory work including in-vivo and time dependent studies.