Microscopic Magnificence
UT Southwestern’s Cryo-Electron Microscopy Facility advances research through imaging technology down to the atomic level
In the 1990s television cartoon series The Magic School Bus, a science teacher, her students, and their class pet – along with the eponymous bus they took on field trips – often shrank to microscopic proportions to explore inside the human body. Their quests, which were meant to teach the fundamentals of biology to elementary school children, animated a goal that researchers have chased for hundreds of years.
By taking a closer look inside the human body, at scales down to the atomic level, researchers believed they could solve fundamental questions about why and how diseases develop, right down to their molecular source, sparking new ideas and therapies that would have vast implications on the future of medicine.
A little less than a decade ago, such ultramicroscopic imaging took a giant leap ahead at UT Southwestern with the opening of the Cryo-Electron Microscopy Facility (CEMF), which houses six cryogenic electron (cryo-EM) microscopes at two locations on the North and South campuses. These highly sought-after instruments, which have superhero names like Titan Krios and Talos Arctica, have vastly broadened the scope of molecules for which researchers are able to determine their 3D structures, accelerating the pace at which they can use this information to better understand diseases and develop new drugs to treat them. Scientists at UT Southwestern and elsewhere have made countless discoveries – placing UTSW among the most respected and in-demand institutions in the world for cryo-EM investigation and creating a significant impact on human health.
What is cryo-electron microscopy?
Microscopy got its start around 1590 in Germany with the invention of the first compound microscopes. For more than three centuries, researchers used variations of this early innovation – which relies on visible light – to image progressively smaller objects. These technologies are still in use today.
But light microscopy has its limits, explained Daniel Stoddard, Ph.D., Assistant Professor of Biophysics and Director of the CEMF: Visible light can only image objects on the order of its wavelength. Like visible light, electrons also have wavelengths, but significantly smaller in scale. With these much smaller wavelengths, electrons can image vastly smaller objects – even at the atomic level – shedding light on the structure of molecules to help researchers better understand their roles in organisms.
“Cryo-EM pushes our understanding of disease from the cellular level to the molecular level,” Dr. Stoddard said.
To accomplish this feat, the right conditions are key, he added. Because biological molecules are inherently mobile in solution, literally freezing them in place is crucial to prevent motion-induced blur. Additionally, since ice crystals can damage biological samples and distort the path of electrons moving through them, a special type of freezing called vitrification is necessary. By plunging a sample into a bath of liquid ethane or propane – itself kept at cryogenic temperatures with liquid nitrogen – residual water in a sample flash-freezes into a glassy state in a thin layer surrounding samples, allowing an electron beam to travel through.
Although scientists have had the ability to utilize cryo-electron microscopy since the 1970s, it wasn’t a suitable imaging technique for defining structures at molecular or atomic resolution for several decades, said Zhe “James” Chen, Ph.D., Professor of Biophysics and co-Director of UTSW’s Structural Biology Lab (SBL).
“The resolution for cryo-EM used to be much lower compared to other structural biology methods such as X-ray crystallography,” Dr. Chen said. “You might be able to see the overall shape of a protein, but getting detailed structural information was out of the question.”
A resolution revolution
Advances in the early 2010s finally made modern cryo-EM possible: The development in both hardware and software sparked a “resolution revolution” that brought cryo-EM resolution down to just a few angstroms, Dr. Chen said. Angstroms are a unit of length equal to one ten-billionth of a meter, or 0.1 nanometers.
Inside the CEMF at UTSW
Today there are more than 60 cryo-EM facilities across the country. CEMF and most others perform single-particle cryo-EM, the workhorse of this technology, by placing a drop of a purified sample containing millions or billions of copies of the same molecule or protein onto a special cryo-EM grid. After vitrification, a cryo-electron microscope bombards the sample with electrons that are collected by a detector. Software then makes sense of the massive amount of data, generating three-dimensional images that allow researchers to visualize the structure of the molecule or protein of interest at atomic-level resolution.
Many structures resolved through cryo-EM, particularly those of greater size and complexity, were beyond reach just a decade ago, Dr. Chen said.
These leaps in cryo-EM earned three scientists, who contributed to the advances, the 2017 Nobel Prize in Chemistry. But even before then, numerous researchers – including many at UTSW – had started turning to this technology to make their own
The vision of cryo-EM
For much of his career, Michael Rosen, Ph.D., Chair and Professor of Biophysics, had largely relied on X-ray crystallography, a technique developed in the early 1900s, to determine the structures of various molecules and proteins. But some materials will not crystallize, a feature necessary to use this technology. Seeing the power of cryo-EM after the resolution revolution, Dr. Rosen and Sandra Schmid, Ph.D., former Chair of Cell Biology at UTSW, decided that the best way to harness the benefits of this new technology was for the University to have its own facility.
“We had a common vision for the future of imaging on our campus,” Dr. Rosen said, “and it was cryo-EM.”
Together, they recruited Daniela Nicastro, Ph.D., now Professor of Cell Biology, to UTSW from Brandeis University. Dr. Stoddard, a graduate student with Dr. Nicastro at the time, came with her. Together, the team and the institution worked to apply for grants to build the CEMF. The first facility, on UTSW’s North Campus, was completed in 2016 with joint funding from UTSW, the Cancer Prevention and Research Institute of Texas (CPRIT), and other sources. A second facility, on South Campus, was built in 2021. A third facility, for performing a related technology called cryo-focused ion beam milling, opened in 2024. The CEMF, which includes all three locations, represents a joint investment of nearly $40 million.
The CEMF also performs cryo-electron tomography, a related technique typically used on larger structures in which the microscope’s stage tilts to capture images of the sample from different angles.
Accelerating discoveries, collaborations
Having CEMF right on campus has been an extraordinary resource for UTSW researchers, accelerating their work and fostering cross-disciplinary collaborations across the institution, said Joan Conaway, Ph.D., Vice Provost, Dean of Basic Research, and Professor of Molecular Biology at UTSW.
When Dr. Conaway arrived at UTSW in 2021, she left a 20-year career at the Stowers Institute for Medical Research in Kansas City, Missouri. There, she and her colleagues used cryo-EM to study various mechanisms involved in transcription, the process in which cells copy DNA into RNA as part of producing proteins. But since they weren’t experts on this technology and there was no resource like CEMF at Stowers, they had to rely on collaborators at other institutions, slowing the pace of research, she said.
“Being able to take your sample and work in an iterative fashion, back and forth with a colleague right here at UTSW, is infinitely better than having to rely on an outside resource,” Dr. Conaway said.
Using the CEMF, UTSW researchers have made many significant breakthroughs. Xiaochen Bai, Ph.D., Associate Professor of Biophysics and Cell Biology and an expert on cryo-EM recruited to UTSW in 2017, solved the structure of the insulin receptor bound to insulin using the technology in 2019.
“Our dream is to develop a better and more affordable insulin accessible to everyone with diabetes,” he said.
Dr. Bai and the Bai Lab are currently working with UTSW researcher Xuewu Zhang, Ph.D., Professor of Pharmacology and Biophysics, who is an expert in structural biology of innate immunity, and with Zhijian “James” Chen, Ph.D., Professor of Molecular Biology and in the Center for the Genetics of Host Defense, who won the prestigious Lasker Award in 2024 for his discovery of the cGAS enzyme. This protein serves as a cellular burglar alarm, Dr. Bai explained, sensing foreign DNA in cells and indirectly activating another protein called Stimulator of Interferon Genes (STING), which launches an immune response. The team is using cryo-EM to learn the structure of STING in both its activated and inactivated state, research that could eventually lead to drugs both to heighten immune responses against cancer and infections and to dampen it to treat autoimmune diseases.
Dr. Rosen said he’s finalizing one of his first studies using cryo-EM, an accomplishment five years in the making. The Rosen Lab studies biomolecular condensates, liquid structures that form separate compartments without membranes in the aqueous environment inside cells, much like oil separates into distinct globules in water. Using cryo-electron tomography, Dr. Rosen and his colleagues are investigating how molecules are spatially arranged in the condensates, findings that could lend insight on their function.
Some other researchers using cryo-EM for their studies at UTSW include:
- Dr. Nicastro, who is shedding light on the 3D structures and functions of organelles and macromolecular complexes, with special emphasis on proteins in the cytoskeleton (interlinking proteins that help cells maintain shape and migrate) and molecular motors (proteins that move materials within cells). Her work is also focused on improving cryo-EM itself.
- Margaret Phillips, Ph.D., Chair and Professor of Biochemistry and Professor of Pharmacology, is using cryo-EM to search for therapeutic targets in the malaria-causing parasite Plasmodium.
- Youxing Jiang, Ph.D., Professor of Physiology and Biophysics, is using cryo-EM to better understand ion channels and membrane transporters, proteins that regulate many biological processes such as the excitation of nerve and muscle cells, the secretion of hormones, and the process by which sensory nerves convert external stimuli into electrical signals in the brain.
Improving cryo-EM
Although significant strides have been made in the use of cryo-EM, Dr. Stoddard said, this pivotal technology still has room for improvement. A substantial goal of biological imaging is being able to see specific objects and processes at high resolution in their native environment inside cells, a feat not possible with traditional cryo-EM.
An important advance toward this goal is cryo-focused ion beam milling, a technique pioneered by Dr. Nicastro, in which researchers carve away portions of larger structures, such as cells, to thin them down and expose items of interest within – for example, specific organelles or protein assemblies. However, Dr. Stoddard noted, this technique destroys parts of the sample that are cut away, and it is still difficult to identify structures of interest during the milling process.
A solution to the where-to-mill problem, he added, is correlative light and electron microscopy (CLEM), a technique in which structures can be tagged with fluorescent molecules inside cells, allowing researchers to spot them more easily. Researchers are also working on improving cryo-EM’s resolution even further.
But as scientists at UTSW and elsewhere tackle these challenges, Dr. Stoddard said, cryo-EM will continue to generate findings that are revolutionizing medicine.
“We hope this technology continues to grow in size and scope, but it’s already affecting people’s lives, transforming how we think about the nature of biological processes, diseases, and drug design,” he said. “It’s everything that medicine has always promised, and it’s happening right now.”
Dr. Bai is a Virginia Murchison Linthicum Scholar in Medical Research.
Dr. Zhijian “James” Chen holds the George L. MacGregor Distinguished Chair in Biomedical Science.
Dr. Conaway holds the Cecil H. Green Distinguished Chair in Cellular and Molecular Biology.
Dr. Jiang holds the Rosewood Corporation Chair in Biomedical Science and is a W.W. Caruth, Jr. Scholar in Biomedical Research.
Dr. Phillips holds The Sam G. Winstead and F. Andrew Bell Distinguished Chair in Biochemistry.
Dr. Rosen holds the Mar Nell and F. Andrew Bell Distinguished Chair in Biochemistry.
Dr. Zhang is a Virginia Murchison Linthicum Scholar in Medical Research.