Supramolecular assembly of collagen fibrils into collagen fiber in fish scales of red seabream, Pagrus major.

Supramolecular assembly of collagen fibrils into collagen fiber and its distribution in fish scales of red seabream, Pagrus major, were investigated. By virtue of Zernike phase-contrast hard X-ray microscopy, it has been firstly observed that collagen fiber consists of helical substructures of collagen fibrils wrapped with incrustation. As it close to the scalar focus (that is, with aging), loosened- and deteriorated-helical assemblies started to be observed with loosing wrapping incrustation, indicative of the distortion of the basic helical assembly. Various distributions and packing arrangements of collagen fibers were observed dependent on subdivisions of fish scale. Freshly growing edge region of fish scale, embedded into fish skin, showed rarely patched and one directionally arranged collagen fibers, in which specifically triple helical assemblies of collagen fibrils were found. On the contrary, relatively aged region of the rostral field close to the scalar focus displayed randomly directed and densely packed collagen fibers, in which loosened- and deteriorated-helical assemblies of collagen fibrils were mostly found. Our results have demonstrated that hard X-ray microscope can be a powerful tool to study in situ internal structure of biological specimens in an atmospheric pressure.

A new method for investigation of the hair shaft: hard X-ray microscopy with a 90-nm spatial resolution.

Various methods have been used to investigate the hair shaft. In the ultrastructural hair field, scanning and transmission electron microscopies are widely used investigative methods, but they have some technical limitations. Recently, X-ray microscopes with sub-micron spatial resolution have emerged as useful instruments because they offer a unique opportunity to observe the interior of an undamaged sample in greater detail. In this report, we examined damaged hair shaft tips using hard X-ray microscopy with a 90 nm spatial resolution. The results of this study suggest that hard X-ray microscopy is an alternative investigative method for hair morphology studies.

X-ray imaging of various biological samples using a phase-contrast hard X-ray microscope.

In this study, we visualized the internal structures of various bio-samples and found the optimum conditions of test samples for the 7 keV hard X-ray microscope of the Pohang light source. From the captured X-ray images, we could observe the intercellular and intracellular structures of dehydrated human cells and mouse tumor tissues without using any staining materials in a spatial resolution better than 100 nm. The metastasized lung tissue, which was several tens of micrometers in thickness, was found to be very well suited to this hard X-ray microscope system, because it is nearly impossible to observe such a nontransparent and thick sample with a high spatial resolution better than 100 nm using any microscopes such as a soft X-ray microscope, an optical microscope, or an electron microscope.

High resolution X-ray phase contrast synchrotron imaging of normal and ligation damaged rat sciatic nerves.

This study was performed to apply synchrotron radiation (SR) imaging to a neuropathologic evaluation technique after treatment of peripheral nerve blocks. A phase contrast synchrotron images of normal and ligation damaged rat sciatic nerve were obtained with an 8 KeV monochromatic beam and 20-mum thick CsI(TI) scintillation crystal. The visual image was magnified using a 20x microscope objective and captured using an analog CCD camera. Obtained images were compared with conventional light microscopic findings from the same nerve samples. By using an edge enhancement effect of phase const with SR, we could easily discriminate each nerve fiber and identify the arrangement of nerve fibers within a whole thickness (about 1 mm in diameter) of peripheral nerve without sectioning and fixation. The composite SR image of a ligation damaged rat sciatic nerve sample showed that the response to nerve injury was different on each side of the site of injury. The SR image of damaged distal lesion showed destruction of neural microarchitecture and typical extensive Wallerian degeneration of nerve fibers as clearly as histologic image. We could get very detailed morphologic data for Wallerian degeneration of nerve fibers by using the SR imaging technique. We believe that the phase contrast synchrotron imaging has great potential as an imaging tool in the bioscience and medical science.

A study of the human hair structure with a Zernike phase contrast X-ray microscope.

We have observed the internal structure of human hair shafts with a transmission Zernike phase contrast hard X-ray microscope. Due to the high spatial resolution and the high contrast of the microscope, we could image scales, macrofibrils, medulla and melanin without staining. The structure of a black hair shaft is compared with that of a white one.

Non-invasive evaluation of hair interior morphology by X-ray microscope.

Lots of trials have been performed to obtain better microscopic images of hair structure. Although scanning electron microscopy (SEM) and transmission electron microscopy (TEM) provide detailed images of hair, artificial processing may modify the original images during sample preparation. To overcome this limitation, we applied newly-developed X-ray microscopy with an 80-100 nm spatial resolution to produce a detailed view of the morphological change of hair interior. This X-ray microscopy permits us to penetrate a hair shaft without any artificial change and also provides precise images of hair interior with fine resolution. We evaluated the interior morphological change of Japanese standard hair tress No. 8 by various treatments such as ultraviolet (UV) irradiation, heating, hair dyeing, decolorizing bleaching agents and permanent waving. Internal morphological images were relatively similar in both heated and UV-irradiated hair. They revealed coarsening of cuticles and dehydration of cortex and medulla. When compared with TEM, X-ray microscopy provides more intact images in the cuticle and may be a useful tool in observing fine cracks of hair cortex. Additionally, X-ray images were intact and not influenced by any processing procedures. In observing the external and internal structure of hair, its resolution seems to be somewhat lesser than TEM and there are also several remaining weaknesses to be improved. Hopefully, forthcoming technology will solve these problems in the near future.

Hard X-ray microscopy with Zernike phase contrast.

Zernike phase contrast has been added to a full-field X-ray microscope with Fresnel zone plates that was in operation at 6.95 keV. The spatial resolution has also been improved by increasing the magnification of the microscope objective looking at the CsI(Tl) scintillation crystal. Cu no. 2000 meshes and a zone plate have been imaged to see the contrast as well as the spatial resolution. A Halo effect coming from the Zernike phase contrast was clearly visible on the images of meshes.

High-resolution X-ray refraction imaging of rat lung and histological correlations.

This study was performed to observe microstructures of the rat lung, using a synchrotron radiation beam and to compare findings with histological observations. X-ray refraction images from ex-vivo ventilating rat lung were obtained with an 8 KeV monochromatic beam and 20-mum thick CsI(Tl) scintillation crystal. The visual image was magnified using a 20x microscope objective and captured using an analog CCD camera. Obtained images were compared with conventional light microscopic findings from the same tissue. Pulmonary microstructures, including alveolar ducts, alveolar sacs, alveoli, alveolar walls, and perialveolar capillary networks were clearly identified with spatial resolution of as much as 1.2 mum and had good correlation with conventional light microscopic findings. The shape of alveoli appeared more round in SR images than in the light microscopic images. The results suggest that X-ray microscopy study of the lung using synchrotron radiation demonstrates the potential for clinically relevant microstructure of lung tissue without sectioning and fixation.

Observations of a human hair shaft with an x-ray microscope.

We observed the internal structures of a human hair shaft using x-ray microscopes with a spatial resolution in the range from a few microns to less than 100 nm. The energy of the x-ray used is 6.95 keV. The Zernike phase contrast together with a spatial resolution better than 100 nm enabled us to see the cuticles of scales, the cortex of macrofibrils and the medulla. All these internal features and more can easily be observed with no sample preparation including staining.