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| Scientists at PSI were able to observe the local collagen structures in an ossicle by scanning it with an X-ray beam. The different colours of the cylinders indicate how strongly the collagen bundles are spatially aligned in a section measuring 20 by 20 by 20 micrometres. Image Credit: © Paul Scherrer Institute PSI/Christian Appel |
Scientific Frontline: "At a Glance" Summary
- Main Discovery: Researchers refined a "tensor tomography" X-ray diffraction technique that simultaneously detects biological structures ranging from nanometers to millimeters, significantly accelerating the imaging process.
- Methodology: The team used a precisely rotated X-ray beam (approx. 20 micrometers wide) to generate millions of interference patterns around two axes, which software then reconstructed into a 3D tomogram.
- Key Statistic: The optimized process reduced the measurement time for a complete tomogram from roughly 24 hours to just over one hour.
- Context: To validate the method, the team imaged the auditory ossicle (anvil) of the ear, successfully mapping the spatial orientation of nanometer-sized collagen fibers crucial for sound transmission.
- Significance: This drastic reduction in scan time makes statistical studies involving hundreds of samples feasible, aiding biomedical research in areas like bone tissue analysis and implant development.
Biological materials are masterpieces created by nature. Bones, for example, are extremely hard, yet at the same time elastic enough to withstand lateral forces without breaking easily. This combination of properties results from their hierarchical structure as composite materials – they combine materials that have different structures on different scales. Human-made composite materials are similar in the way they are made up. In reinforced concrete, for example, the concrete component, consisting of cement and sand, can withstand high pressure, while a steel mesh provides high tensile strength and transverse stability.
Until now, examining such biological materials in detail has required the use of several different instruments, such as electron microscopes or classic light microscopes. However, scientists at the PSI Center for Photon Science have now refined an X-ray diffraction technique that was developed at the institute ten years ago, allowing it to be used to characterize materials on scales from nanometers to millimeters simultaneously and much faster than before. A complete scan now only takes about an hour, instead of a whole day.
To demonstrate the efficiency of their method, the researchers used the Swiss Light Source SLS to reveal the alignment of collagen fibers in a human ossicle known as the incus, or anvil. Collagen fibers are thread-like protein structures that provide tensile strength and elasticity to bones. “In doing so, we have taken the leap from a scientific method to a practical technique,” says Christian Appel, postdoctoral researcher and first author of the study. The results have now been published in the journal Small Methods as its cover story. In future, this method could be valuable in areas such as the study of complex tissue, the analysis of bone diseases, and the optimization of implant designs.
Spatial resolution via rasterization
One way of using X-rays is for classic imaging of the kind often seen in hospitals. Those rays that are not absorbed by the tissue pass through to the other side, where they can be viewed as a light and dark image, a radiographic projection. Another approach is to use X-rays like those produced at the SLS to reveal crystal structures at the nanometer scale, exploiting a phenomenon known as interference.
Interference occurs when the electromagnetic waves of the X-rays are scattered by the regularly arranged atomic layers in the crystal and made to overlap. Depending on the direction in which they are scattered, the waves travel different distances to the X-ray camera, where they are recorded, arriving there with different phases – that is to say, the waves are slightly shifted with respect to each other. Consequently, they either reinforce or cancel each other out. The resulting interference pattern can then be used to determine the crystal structure and its orientation in space.
The bone’s ultra-fine collagen fibrils are only visible on the nanometer scale, while coarser tissue structures are visible in the micro- to millimeter range. Ten years ago, researchers led by Marianne Liebi at PSI developed a special technique known as tensor tomography, which can record both length scales simultaneously. The sample to be analyzed is rotated step by step, with high precision, around two axes. At each step, an X-ray beam with a width of only around twenty micrometers produces an interference pattern, which is recorded by a camera. “This rasterization allows information about the local crystal properties to be recorded little by little,” explains Marianne Liebi, who is also lead author of the new study.
A computer program then calculates a three-dimensional image of the entire sample, known as a tomogram, from millions of interference patterns. Originally, the whole process used to take up to a day, which made statistical studies involving hundreds of different samples – as are necessary in modern biomedical research – almost impossible. “We have now refined the method so that we can record a complete tomogram in just over an hour,” says Meitian Wang, beamline scientist and co-author of the study. Thanks to a broad-based collaboration involving several research groups at PSI, it was possible to improve both the scanning technique, in which the object must be positioned extremely precisely and rotated in tiny steps, and the computer software, which calculates the finished tomogram from the individual interference patterns.
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Marianne Liebi, Meitian Wang and Christian Appel (from left to right) at the Swiss Light Source SLS. It was here that they developed a new X-ray technique that allows biological structures to be recorded at scales ranging from nanometres to millimetres in record time.
Photo Credit: © Paul Scherrer Institute PSI/Mahir Dzambegovic
Collagen structures in bone
“In order to test our improved method, we needed an interesting sample,” says Appel. In collaboration with researchers from the Lausanne University Hospital, they chose a tiny bone found in the ear, known as the anvil. It is just a few millimeters across but crucial to hearing. The anvil transmits sound vibrations from the eardrum to the inner ear. If the anvil is damaged, for example by chronic inflammation of the middle ear, it sometimes becomes necessary to replace part of it with a prosthesis. To do this, doctors would like to know exactly what the inside of the anvil looks like.
The collagen structures and their spatial orientation in the bone are particularly instructive. The protein collagen fulfils the same function in bones that steel mesh does in reinforced concrete: it provides stability and at the same time elasticity. The orientation of the collagen fibers can help to determine the best way of attaching a prosthesis. Using the interference patterns recorded during the X-ray scan, the computer program calculated the average alignment of the collagen fibers in many tiny sections of the anvil measuring just 20 by 20 by 20 micrometers. For comparison, the diameter of human hair is around 50 micrometers.
The new SLS will allow the scientists to significantly improve this already very high resolution even further in the future. The width of the X-ray beam will shrink to just a few micrometers, and the higher X-ray flux will allow routine measurements to be conducted more quickly. “Combining a higher resolution with a higher speed of the measurement opens up completely new possibilities for tensor tomography, especially in biomedical applications,” says Christian Appel.
Published in journal: Small Methods
Authors: Christian Appel, Margaux Schmeltz, Irene Rodriguez-Fernandez, Lukas Anschuetz, Leonard C. Nielsen, Ezequiel Panepucci, Tomislav Marijolovic, Klaus Wakonig, Aleksandra Ivanovic, Anne Bonnin, Filip Leonarski, Justyna Wojdyla, Takashi Tomizaki, Manuel Guizar-Sicairos, Kate Smith, John H. Beale, Wayne Glettig, Katherine E. McAuley, Oliver Bunk, Meitian Wang, and Marianne Liebi
Source/Credit: Paul Scherrer Institute
Reference Number: ms011226_01