Scientists at the National Synchrotron Light Source II (NSLS-II) – a US Department of Energy (DOE) of the US Department of Science (DOE) at the US Brookhaven National Laboratory – use ultra-bright X-rays individual bacteria with higher concentration imaged spatial resolution than ever before. Their work, published in Scientific Reports shows an X-ray imaging technique called X-ray fluorescence microscopy (XRF) as an effective approach for generating 3-D images of small biological samples.
"For the first time, we used nanoscale X-ray fluorescence filters to image bacteria up to cell membrane dissolution," said Lisa Miller, NSLS-II scientist and co-author of the paper. "The imaging of cells at the membrane level is critical to understanding the role of the cell in various diseases and the development of advanced medical treatments."
The record breaking resolution of the X-ray images was made possible by the advanced capabilities of the Hard X-ray Nanoprobe Beamline (HXN), an experimental station at NSLS-II with novel nanofocusing optics and exceptional stability.
"HXN is the first XRF beamline to produce a 3-D image with this type of resolution," Miller said
While other imaging techniques, such as electron microscopy, map the structure of a very high-resolution cell membrane These methods can not provide chemical information about the cell. At HXN, researchers were able to create three-dimensional chemical maps of their samples to find out where trace elements are found throughout the cell.
"At HXN, we take a sample of a sample at an angle and rotate the sample to the next take another image and so on," said Tiffany Victor, lead author of the study and NSLS-II scientist. "Each image shows the chemical profile of the sample in that orientation, and then we can put those profiles together to create a 3D image."
Miller added, "Getting an XRF-3-D image is similar to a regular image X-rays can be obtained in the doctor's office for a CT scan."
The images generated by HXN showed two Trace elements, calcium and zinc, had unique spatial distributions in the bacterial cell.
Zinc is associated with the ribosomes in the bacteria, "said Victor." Bacteria do not have many cellular organelles, unlike eukaryotic (complex) cells, which have mitochondria, a nucleus, and many other organelles. So it's not the most exciting sample for a picture, but it's a nice model system that demonstrates the imaging technique excellently.
Yong Chu, chief beamline scientist at HXN, says the imaging technique is also applicable to many other research areas.
"This chemical 3D imaging or fluorescence nanotome imaging technique is gaining popularity in other scientific fields." Chu said, "For example, we can imagine how the internal structure of a battery changes as it is charged and discharged."
In addition to breaking the technical barriers to X-ray resolution with this technique, researchers developed one new method for imaging bacteria at room temperature during X-ray measurements.
"RFA imaging should ideally be performed on frozen biological samples that are cryopreserved to prevent radiation damage and to provide a more physiologically relevant understanding of cellular processes," said Victor. "Because of the cramped space In the HXN sample chamber, we were unable to examine the sample with one cryostage. Instead, we embedded the cells in small sodium chloride crystals and decanted the cells at room temperature. The sodium chloride crystals retained the rod in a similar form of the cells, making it easier to locate the cells, thereby shortening the duration of our experiments. The researchers say that the effectiveness of X-ray imaging technology and the sample preparation method are demonstrated was the first step in a larger scale nanoscale imaging of trace elements in other biological cells, and the team is particularly interested in the role of copper in the death of copper Neurons in Alzheimer's Disease.
"Trace elements such as iron, copper and zinc are nutritionally essential but may also play a role in disease," said Miller. "We want to understand the subcellular location and function of metal-containing proteins in the disease process. to contribute to the development of effective therapies. "
The work was supported by the DOE's Office of Science, the National Institutes of Health, and the National Science Foundation.