Tagged: Electron Microscopy Toggle Comment Threads | Keyboard Shortcuts

  • richardmitnick 11:53 am on July 29, 2017 Permalink | Reply
    Tags: 3D structure of human chromatin, , ChromEMT, , Electron Microscopy, , , ,   

    From Salk: “Salk scientists solve longstanding biological mystery of DNA organization” 

    Salk Institute bloc

    Salk Institute for Biological Studies

    July 27, 2017

    Stretched out, the DNA from all the cells in our body would reach Pluto. So how does each tiny cell pack a two-meter length of DNA into its nucleus, which is just one-thousandth of a millimeter across?

    The answer to this daunting biological riddle is central to understanding how the three-dimensional organization of DNA in the nucleus influences our biology, from how our genome orchestrates our cellular activity to how genes are passed from parents to children.

    Now, scientists at the Salk Institute and the University of California, San Diego, have for the first time provided an unprecedented view of the 3D structure of human chromatin—the combination of DNA and proteins—in the nucleus of living human cells.

    In the tour de force study, described in Science on July 27, 2017, the Salk researchers identified a novel DNA dye that, when paired with advanced microscopy in a combined technology called ChromEMT, allows highly detailed visualization of chromatin structure in cells in the resting and mitotic (dividing) stages. By revealing nuclear chromatin structure in living cells, the work may help rewrite the textbook model of DNA organization and even change how we approach treatments for disease.

    “One of the most intractable challenges in biology is to discover the higher-order structure of DNA in the nucleus and how is this linked to its functions in the genome,” says Salk Associate Professor Clodagh O’Shea, a Howard Hughes Medical Institute Faculty Scholar and senior author of the paper. “It is of eminent importance, for this is the biologically relevant structure of DNA that determines both gene function and activity.”

    A new technique enables 3D visualization of chromatin (DNA plus associated proteins) structure and organization within a cell nucleus (purple, bottom left) by painting the chromatin with a metal cast and imaging it with electron microscopy (EM). The middle block shows the captured EM image data, the front block illustrates the chromatin organization from the EM data, and the rear block shows the contour lines of chromatin density from sparse (cyan and green) to dense (orange and red). Credit: Salk Institute.

    Ever since Francis Crick and James Watson determined the primary structure of DNA to be a double helix, scientists have wondered how DNA is further organized to allow its entire length to pack into the nucleus such that the cell’s copying machinery can access it at different points in the cell’s cycle of activity. X-rays and microscopy showed that the primary level of chromatin organization involves 147 bases of DNA spooling around proteins to form particles approximately 11 nanometers (nm) in diameter called nucleosomes. These nucleosome “beads on a string” are then thought to fold into discrete fibers of increasing diameter (30, 120, 320 nm etc.), until they form chromosomes. The problem is, no one has seen chromatin in these discrete intermediate sizes in cells that have not been broken apart and had their DNA harshly processed, so the textbook model of chromatin’s hierarchical higher-order organization in intact cells has remained unverified.

    To overcome the problem of visualizing chromatin in an intact nucleus, O’Shea’s team screened a number of candidate dyes, eventually finding one that could be precisely manipulated with light to undergo a complex series of chemical reactions that would essentially “paint” the surface of DNA with a metal so that its local structure and 3D polymer organization could be imaged in a living cell. The team partnered with UC San Diego professor and microscopy expert Mark Ellisman, one of the paper’s coauthors, to exploit an advanced form of electron microscopy that tilts samples in an electron beam enabling their 3D structure to be reconstructed. By combining their chromatin dye with electron-microscope tomography, they created ChromEMT.

    The team used ChromEMT to image and measure chromatin in resting human cells and during cell division when DNA is compacted into its most dense form—the 23 pairs of mitotic chromosomes that are the iconic image of the human genome. Surprisingly, they did not see any of the higher-order structures of the textbook model anywhere.

    From left: Horng Ou and Clodagh O’Shea. Credit: Salk Institute.

    “The textbook model is a cartoon illustration for a reason,” says Horng Ou, a Salk research associate and the paper’s first author. “Chromatin that has been extracted from the nucleus and subjected to processing in vitro—in test tubes—may not look like chromatin in an intact cell, so it is tremendously important to be able to see it in vivo.”

    What O’Shea’s team saw, in both resting and dividing cells, was chromatin whose “beads on a string” did not form any higher-order structure like the theorized 30 or 120 or 320 nanometers. Instead, it formed a semi-flexible chain, which they painstakingly measured as varying continuously along its length between just 5 and 24 nanometers, bending and flexing to achieve different levels of compaction. This suggests that it is chromatin’s packing density, and not some higher-order structure, that determines which areas of the genome are active and which are suppressed.

    With their 3D microscopy reconstructions, the team was able to move through a 250 nm x 1000 nm x 1000 nm volume of chromatin’s twists and turns, and envision how a large molecule like RNA polymerase, which transcribes (copies) DNA, might be directed by chromatin’s variable packing density, like a video game aircraft flying through a series of canyons, to a particular spot in the genome. Besides potentially upending the textbook model of DNA organization, the team’s results suggest that controlling access to chromatin could be a useful approach to preventing, diagnosing and treating diseases such as cancer.

    “We show that chromatin does not need to form discrete higher-order structures to fit in the nucleus,” adds O’Shea. “It’s the packing density that could change and limit the accessibility of chromatin, providing a local and global structural basis through which different combinations of DNA sequences, nucleosome variations and modifications could be integrated in the nucleus to exquisitely fine-tune the functional activity and accessibility of our genomes.”

    Future work will examine whether chromatin’s structure is universal among cell types or even among organisms.

    Other authors included Sébastien Phan, Thomas Deerinck and Andrea Thor of the UC San Diego.

    The work was largely funded by the W. M. Keck Foundation, the NIH 4D Nucleome Roadmap Initiative and the Howard Hughes Medical Institute, with additional support from the William Scandling Trust, the Price Family Foundation and the Leona M. and Harry B. Helmsley Charitable Trust.

    See the full article here .

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition

    Salk Institute Campus

    Every cure has a starting point. Like Dr. Jonas Salk when he conquered polio, Salk scientists are dedicated to innovative biological research. Exploring the molecular basis of diseases makes curing them more likely. In an outstanding and unique environment we gather the foremost scientific minds in the world and give them the freedom to work collaboratively and think creatively. For over 50 years this wide-ranging scientific inquiry has yielded life-changing discoveries impacting human health. We are home to Nobel Laureates and members of the National Academy of Sciences who train and mentor the next generation of international scientists. We lead biological research. We prize discovery. Salk is where cures begin.

  • richardmitnick 9:28 am on July 24, 2017 Permalink | Reply
    Tags: , , Electron Microscopy, , Native mass spectrometry, Signaling islands in cells: targets for precision drug design,   

    From U Washington: “Signaling islands in cells: targets for precision drug design” 

    U Washington

    University of Washington

    Leila Gray

    A critical component of the cell signaling system, anchored protein kinase A, has some flexible molecular parts, allowing it to both contract and stretch, with floppy arms that can reach out to find appropriate targets. John Scott Lab.

    Research results reported in the journal Science overturn long-held views on a basic messaging system within living cells.

    The findings suggest new approaches to designing precisely targeted drugs for cancer and other serious diseases.

    Dr. John D. Scott, professor and chair of pharmacology at the University of Washington School of Medicine and a Howard Hughes Medical Institute Investigator, along with Dr. F. Donelson Smith of the UW and HHMI, led this study, which also involved Drs. Claire and Patrick Eyers and their group at the University of Liverpool. Visit the Scott lab web site, Cell Signaling in Space and Time.

    The researchers explained that key cellular communication machinery is more regionally constrained inside the cell than was previously thought. Communication via this vital system is akin to social networking on your Snapchat account.

    Within a cell, the precise positioning of such messaging components allows hormones, the body’s chief chemical communicators, to transmit information to exact places inside the cell. Accurate and very local activation of the enzyme that Scott and his group study helps assure a correct response occurs in the right place and at the right time.

    “The inside of a cell is like a crowded city,” said Scott, “It is a place of construction and tearing down, goods being transported and trash being recycled, countless messages, (such as the ones we have discovered), assembly lines flowing, and packages moving. Strategically switching on signaling enzyme islands allows these biochemical activities to keep the cell alive and is important to protect against the onset of chronic diseases such as diabetes, heart disease and certain cancers.”

    Advances in electron microscopy and native mass spectrometry enabled the researchers to determine that a critical component of the signaling system, anchored protein kinase A, remains intact during activation. Parts of the molecule are flexible, allowing it to both contract and stretch, with floppy arms that can reach out to find appropriate targets.

    Still, where the molecule performs its act, space is tight. The distance is, in fact, about the width of two proteins inside the cell.

    Green, circled area show where the enzyme in the signalling study is active in mitochondria, the powerhouses of living cells. John D. Scott.

    “We realize that in designing drugs to reach such targets that they will have to work within very narrow confines, ” Scott said.

    One of his group’s collective goals is figuring out how to deliver precision drugs to the right address within this teeming cytoplasmic metropolis.

    “Insulating the signal so that the drug effect can’t happen elsewhere in the cell is an equally important aspect of drug development because it could greatly reduce side effects,” Scott said.

    An effort to take this idea of precision medicine a step further is part of the Institute for Targeted Therapeutics at UW Medicine in Seattle. The institute is being set up by Scott and his colleagues in the UW Department of Pharmacology.

    The scientists are collaborating with cancer researchers to better understand the molecular causes — and possible future treatments — for a certain liver malignancy. This particular liver cancer arises from a mutation that produces an abnormal form of the enzyme that is the topic of this current work, protein kinase A, and alters the enzyme’s role in cell signaling.

    Other advances that gave the researchers a clearer view of the signaling mechanisms reported in Science include CRISPR gene editing, live-cell imaging techniques, and more powerful ways to look at all components of a protein complex.

    See the full article here .

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition

    The University of Washington is one of the world’s preeminent public universities. Our impact on individuals, on our region, and on the world is profound — whether we are launching young people into a boundless future or confronting the grand challenges of our time through undaunted research and scholarship. Ranked number 10 in the world in Shanghai Jiao Tong University rankings and educating more than 54,000 students annually, our students and faculty work together to turn ideas into impact and in the process transform lives and our world. For more about our impact on the world, every day.
    So what defines us —the students, faculty and community members at the University of Washington? Above all, it’s our belief in possibility and our unshakable optimism. It’s a connection to others, both near and far. It’s a hunger that pushes us to tackle challenges and pursue progress. It’s the conviction that together we can create a world of good. Join us on the journey.

  • richardmitnick 2:52 pm on April 3, 2017 Permalink | Reply
    Tags: , , Electron Microscopy   

    From Cornell: “New electron microscope sees more than an image” 

    Cornell Bloc

    Cornell University

    March 30, 2017
    Bill Steele

    Sol Gruner, left, professor of physics, and David Muller, professor of applied and engineering physics. Chris Kitchen/University Photography

    . Their electron microscope pixel array detector (EMPAD) yields not just an image, but a wealth of information about the electrons that create the image and, from that, more about the structure of the sample.

    “We can extract local strains, tilts, rotations, polarity and even electric and magnetic fields,” explained David Muller, professor of applied and engineering physics, who developed the new device with Sol Gruner, professor of physics, and members of their research groups.

    Cornell’s Center for Technology Licensing (CTL) has licensed the invention to FEI, a leading manufacturer of electron microscopes (a division of Thermo Fisher Scientific, which supplies products and services for the life sciences through several brands). FEI expects to complete the commercialization of the design and offer the detector for new and retrofitted electron microscopes this year.

    “It’s mind-boggling to contemplate what researchers around the world will discover through this match of Cornell’s deep expertise in detector science with market leader Thermo Fisher Scientific,” said Patrick Govang, technology licensing officer at CTL.

    The scientists described their work in the February 2016 issue of the journal Microscopy and Microanalysis.

    In the usual scanning transmission electron microscope (STEM), a narrow beam of electrons is fired down through a sample, scanning back and forth to produce an image. A detector underneath reads the varying intensity of electrons coming through and sends a signal that draws an image on a computer screen.

    The EMPAD that replaces the usual detector is made up of a 128×128 array of electron-sensitive pixels, each 150 microns (millionths of a meter) square, bonded to an integrated circuit that reads out the signals – somewhat like the array of light-sensitive pixels in the sensor in a digital camera, but not to form an image. Its purpose is to detect the angles at which electrons emerge, as each electron hits a different pixel. The EMPAD is a spinoff of X-ray detectors the physicists have built for X-ray crystallography work at the Cornell High Energy Synchrotron Source (CHESS), and it can work in a similar way to reveal the atomic structure of a sample.

    Combined with the focused beam of the electron microscope, the detector allows researchers to build up a “four-dimensional” map of both position and momentum of the electrons as they pass through a sample to reveal the atomic structure and forces inside. The EMPAD is unusual in its speed, sensitivity and wide range of intensities it can record – from detecting a single electron to intense beams containing hundreds of thousands or even a million electrons.

    “It would be like taking a photograph of a sunset that showed both details on the surface of the sun and the details of darkest shadows,” Muller explained.

    See the full article here .

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition
    Once called “the first American university” by educational historian Frederick Rudolph, Cornell University represents a distinctive mix of eminent scholarship and democratic ideals. Adding practical subjects to the classics and admitting qualified students regardless of nationality, race, social circumstance, gender, or religion was quite a departure when Cornell was founded in 1865.

    Today’s Cornell reflects this heritage of egalitarian excellence. It is home to the nation’s first colleges devoted to hotel administration, industrial and labor relations, and veterinary medicine. Both a private university and the land-grant institution of New York State, Cornell University is the most educationally diverse member of the Ivy League.

    On the Ithaca campus alone nearly 20,000 students representing every state and 120 countries choose from among 4,000 courses in 11 undergraduate, graduate, and professional schools. Many undergraduates participate in a wide range of interdisciplinary programs, play meaningful roles in original research, and study in Cornell programs in Washington, New York City, and the world over.

  • richardmitnick 9:22 am on July 18, 2016 Permalink | Reply
    Tags: A glimpse inside the atom, Electron Microscopy, Electron orbitals, TU Wein   

    From TU Wein: “A glimpse inside the atom” 

    Techniche Universitat Wein (Vienna)

    Techniche Universität Wein (Vienna)

    Florian Aigner

    Further information:
    Dr. Stefan Löffler
    Service-Einrichtung für Transmissions-Elektronenmikroskopie (USTEM)
    TU Wien (Vienna)
    Wiedner Hauptstraße 8, 1040 Vienna

    Prof. Peter Schattschneider
    Institute of Solid State Physics
    TU Wien (Vienna)
    Wiedner Hauptstraße 8, 1040 Vienna

    Using electron microscopes, it is possible to image individual atoms. Scientists at TU Wien have calculated how it is possible to look inside the atom to image individual electron orbitals.

    Atomic orbitals of carbon atoms in graphene

    Peter Schattschneider, Johannes Bernardi, Stefan Löffler

    An electron microscope can’t just snap a photo like a mobile phone camera can. The ability of an electron microscope to image a structure – and how successful this imaging will be – depends on how well you understand the structure. Complex physics calculations are often needed to make full use of the potential of electron microscopy. An international research team led by TU Wien’s Prof. Peter Schattschneider set out to analyse the opportunities offered by EFTEM, that is energy-filtered transmission electron microscopy. The team demonstrated numerically that under certain conditions, it is possible to obtain clear images of the orbital of each individual electron within an atom. Electron microscopy can therefore be used to penetrate down to the subatomic level – experiments in this area are already planned. The study has now been published in the physics journal Physical Review Letters.

    In search of the electron orbital

    We often think of atomic electrons as little spheres that circle around the nucleus of the atom like tiny planets around a sun. This image is barely reflected in reality, however. The laws of quantum physics state that the position of an electron cannot be clearly defined at any given point in time. The electron is effectively smeared across an area close to the nucleus. The area that could contain the electron is called the orbital. Although it has been possible to calculate the shape of these orbitals for a long time, efforts to image them with electron microscopes have been unsuccessful to date.

    “We have calculated how we might have a chance of visualising orbitals with an electron microscope”, says Stefan Löffler from the University Service Centre for Transmission Electron Microscopy (USTEM) at TU Wien. “Graphene, which is made of just one single layer of carbon atoms, is an excellent candidate for this task. The electron ray is able to pass easily through the graphene with hardly any elastic scattering. An image of the graphene structure can be created with these electrons.”

    Researchers have been aware of the principle of “energy-filtered transmission electron microscopy” (EFTEM) for some time. EFTEM can be used to create quite specific visualisations of certain kinds of atoms whilst blocking out the others. For this reason, it is often used today to analyse the chemical composition of microscopic samples. “The electrons shot through the sample can excite the sample’s atoms”, explains Stefan Löffler. “This costs energy, so when the electrons emerging emerge from the sample, they are slower than when they entered it. This velocity and energy change is characteristic for certain excitations of electron orbitals within the sample.”

    After the electrons have passed through the sample, a magnetic field sorts the electrons by energy. “A filter is used to block out electrons that aren’t of interest: the recorded image contains only those electrons that carry the desired information.”

    Defects can be helpful

    The team used simulations to investigate how this technique could help reach a turning point in the study of electron orbitals. While doing so, they discovered something that actually facilitated the imaging of individual orbitals: “The symmetry of the graphene has to be broken”, says Stefan. “If, for instance, there is a hole in the graphene structure, the atoms right beside this hole have a slightly different electronic structure, making it possible to image the orbitals of these atoms. The same thing can happen if a nitrogen atom rather than a carbon atom is found somewhere in the graphene. When doing this, it’s important to focus on the electrons found within a narrow and precise energy window, minimise certain aberrations of the electromagnetic lens and, last but not least, use a first-rate electron microscope.” All of these issues can be overcome, however, as the research group’s calculations show.

    The Humboldt-Universität zu Berlin, the Universität Ulm, and McMaster University in Canada also worked alongside the TU Wien on the study in a joint FWF-DFG project (“Towards orbital mapping”, I543-N20) and a FWF Erwin-Schrödinger project (“EELS at interfaces”, J3732-N27). Ulm is currently developing a new, high-performance transmission electron microscope that will be used to put these ideas into practice in the near future. Initial results have already exceeded expectations.

    Science paper:
    Mapping Atomic Orbitals with the Transmission Electron Microscope: Images of Defective Graphene Predicted from First-Principles Theory

    See the full article here .

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition

    Techniche Universitat Wein (Vienna) campus

    Our mission is “technology for people”. Through our research we “develop scientific excellence”,
    through our teaching we “enhance comprehensive competence”.

    TU Wien (TUW) is located in the heart of Europe, in a cosmopolitan city of great cultural diversity. For nearly 200 years, TU Wien has been a place of research, teaching and learning in the service of progress. TU Wien is among the most successful technical universities in Europe and is Austria’s largest scientific-technical research and educational institution.

  • richardmitnick 4:24 pm on February 29, 2016 Permalink | Reply
    Tags: , Electron Microscopy,   

    From LBL: “New Form of Electron-beam Imaging Can See Elements that are ‘Invisible’ to Common Methods” 

    Berkeley Logo

    Berkeley Lab

    February 29, 2016
    Glenn Roberts Jr. 510-486-5582

    Electrons can extend our view of microscopic objects well beyond what’s possible with visible light—all the way to the atomic scale. A popular method in electron microscopy for looking at tough, resilient materials in atomic detail is called STEM, or scanning transmission electron microscopy, but the highly focused beam of electrons used in STEM can also easily destroy delicate samples.

    This is why using electrons to image biological or other organic compounds, such as chemical mixes that include lithium—a light metal that is a popular element in next-generation battery research—requires a very low electron dose.

    Scientists at the Department of Energy’s Lawrence Berkeley National Laboratory (Berkeley Lab) have developed a new imaging technique, tested on samples of nanoscale gold and carbon, that greatly improves images of light elements using fewer electrons.

    The newly demonstrated technique, dubbed MIDI-STEM, for matched illumination and detector interferometry STEM, combines STEM with an optical device called a phase plate that modifies the alternating peak-to-trough, wave-like properties (called the phase) of the electron beam.

    In MIDI-STEM (right), developed at Berkeley Lab, an electron beam travels through a ringed “phase plate,” producing a high-resolution image (bottom right) that provides details about a sample containing a heavy element (gold) and light element (carbon). Details about the carbon are missing in an image (bottom left) of the sample using a conventional electron imaging technique (ADF-STEM). (Colin Ophus/Berkeley Lab, Nature Communications: 10.1038/ncomms10719)

    This phase plate modifies the electron beam in a way that allows subtle changes in a material to be measured, even revealing materials that would be invisible in traditional STEM imaging.

    Another electron-based method, which researchers use to determine the detailed structure of delicate, frozen biological samples, is called cryo-electron microscopy, or cryo-EM. While single-particle cryo-EM is a powerful tool—it was named as science journal Nature’s 2015 Method of the Year—it typically requires taking an average over many identical samples to be effective. Cryo-EM is generally not useful for studying samples with a mixture of heavy elements (for example, most types of metals) and light elements like oxygen and carbon.

    “The MIDI-STEM method provides hope for seeing structures with a mixture of heavy and light elements, even when they are bunched closely together,” said Colin Ophus, a project scientist at Berkeley Lab’s Molecular Foundry and lead author of a study, published Feb. 29 in Nature Communications, that details this method.

    If you take a heavy-element nanoparticle and add molecules to give it a specific function, conventional techniques don’t provide an easy, clear way to see the areas where the nanoparticle and added molecules meet.

    “How are they aligned? How are they oriented?” Ophus asked. “There are so many questions about these systems, and because there wasn’t a way to see them, we couldn’t directly answer them.”

    While traditional STEM is effective for “hard” samples that can stand up to intense electron beams, and cryo-EM can image biological samples, “We can do both at once” with the MIDI-STEM technique, said Peter Ercius, a Berkeley Lab staff scientist at the Molecular Foundry and co-author of the study.

    The phase plate in the MIDI-STEM technique allows a direct measure of the phase of electrons that are weakly scattered as they interact with light elements in the sample. These measurements are then used to construct so-called phase-contrast images of the elements. Without this phase information, the high-resolution images of these elements would not be possible.

    This animated representation shows a Berkeley Lab-developed technique called MIDI-STEM (at right) and conventional STEM (at left) that does not use a ringed object called a phase plate. In MIDI-STEM, an interference pattern (bottom right) introduced by the phase plate (top right) interacts with the electron beam before it travels through a sample (the blue wave in the center). As the phase of the sample (the distance between the peaks and valleys of the blue wave) changes, the electrons passing through the sample are affected and can be measured as a pattern (bottom right). (Colin Ophus/Berkeley Lab)

    In this study, the researchers combined phase plate technology with one of the world’s highest resolution STEMs, at Berkeley Lab’s Molecular Foundry, and a high-speed electron detector.

    They produced images of samples of crystalline gold nanoparticles, which measured several nanometers across, and the superthin film of amorphous carbon that the particles sat on. They also performed computer simulations that validated what they saw in the experiment.

    The phase plate technology was developed as part of a Berkeley Lab Laboratory Directed Research and Development grant in collaboration with Ben McMorran at University of Oregon.

    The MIDI-STEM technique could prove particularly useful for directly viewing nanoscale objects with a mixture of heavy and light materials, such as some battery and energy-harvesting materials, that are otherwise difficult to view together at atomic resolution.

    It also might be useful in revealing new details about important two-dimensional proteins, called S-layer proteins, that could serve as foundations for engineered nanostructures but are challenging to study in atomic detail using other techniques.

    In the future, a faster, more sensitive electron detector could allow researchers to study even more delicate samples at improved resolution by exposing them to fewer electrons per image.

    “If you can lower the electron dose you can tilt beam-sensitive samples into many orientations and reconstruct the sample in 3-D, like a medical CT scan. There are also data issues that need to be addressed,” Ercius said, as faster detectors will generate huge amounts of data. Another goal is to make the technique more “plug-and-play,” so it is broadly accessible to other scientists.

    Berkeley Lab’s Molecular Foundry is a DOE Office of Science User Facility. Researchers from the University of Oregon, Gatan Inc. and Ulm University in Germany also participated in the study.

    See the full article here .

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition

    A U.S. Department of Energy National Laboratory Operated by the University of California

    University of California Seal

    DOE Seal

  • richardmitnick 2:23 pm on May 14, 2015 Permalink | Reply
    Tags: , Electron Microscopy,   

    From LBL: “CLAIRE Brings Electron Microscopy to Soft Materials” 

    Berkeley Logo

    Berkeley Lab

    May 14, 2015
    Lynn Yarris (510) 486-5375

    Berkeley Researchers Develop Breakthrough Technique for Non-invasive Nano-scale Imaging

    CLAIRE image of Al nanostructures with an inset that shows a cluster of six Al nanostructures.

    Soft matter encompasses a broad swath of materials, including liquids, polymers, gels, foam and – most importantly – biomolecules. At the heart of soft materials, governing their overall properties and capabilities, are the interactions of nano-sized components. Observing the dynamics behind these interactions is critical to understanding key biological processes, such as protein crystallization and metabolism, and could help accelerate the development of important new technologies, such as artificial photosynthesis or high-efficiency photovoltaic cells. Observing these dynamics at sufficient resolution has been a major challenge, but this challenge is now being met with a new non-invasive nanoscale imaging technique that goes by the acronym of CLAIRE.

    CLAIRE stands for “cathodoluminescence activated imaging by resonant energy transfer.” Invented by researchers with the U.S. Department of Energy (DOE)’s Lawrence Berkeley National Laboratory (Berkeley Lab) and the University of California (UC) Berkeley, CLAIRE extends the incredible resolution of electron microscopy to the dynamic imaging of soft matter.

    “Traditional electron microscopy damages soft materials and has therefore mainly been used to provide topographical or compositional information about robust inorganic solids or fixed sections of biological specimens,” says chemist Naomi Ginsberg, who leads CLAIRE’s development. “CLAIRE allows us to convert electron microscopy into a new non-invasive imaging modality for studying soft materials and providing spectrally specific information about them on the nanoscale.”

    Naomi Ginsberg

    Ginsberg holds appointments with Berkeley Lab’s Physical Biosciences Division and its Materials Sciences Division, as well as UC Berkeley’s departments of chemistry and physics. She is also a member of the Kavli Energy NanoScience Institute (Kavli-ENSI) at Berkeley. She and her research group recently demonstrated CLAIRE’s imaging capabilities by applying the technique to aluminum nanostructures and polymer films that could not have been directly imaged with electron microscopy.

    “What microscopic defects in molecular solids give rise to their functional optical and electronic properties? By what potentially controllable process do such solids form from their individual microscopic components, initially in the solution phase? The answers require observing the dynamics of electronic excitations or of molecules themselves as they explore spatially heterogeneous landscapes in condensed phase systems,” Ginsberg says. “In our demonstration, we obtained optical images of aluminum nanostructures with 46 nanometer resolution, then validated the non-invasiveness of CLAIRE by imaging a conjugated polymer film. The high resolution, speed and non-invasiveness we demonstrated with CLAIRE positions us to transform our current understanding of key biomolecular interactions.”

    CLAIRE imaging chip consists of a YAlO3:Ce scintillator film supported by LaAlO3 and SrTiO3 buffer layers and a Si frame. Al nanostructures embedded in SiO2 are positioned below and directly against the scintillator film. ProTEK B3 serves as a protective layer for etching.

    CLAIRE works by essentially combining the best attributes of optical and scanning electron microscopy into a single imaging platform. Scanning electron microscopes use beams of electrons rather than light for illumination and magnification. With much shorter wavelengths than photons of visible light, electron beams can be used to observe objects hundreds of times smaller than those that can be resolved with an optical microscope. However, these electron beams destroy most forms of soft matter and are incapable of spectrally specific molecular excitation.

    Ginsberg and her colleagues get around these problems by employing a process called “cathodoluminescence,” in which an ultrathin scintillating film, about 20 nanometers thick, composed of cerium-doped yttrium aluminum perovskite, is inserted between the electron beam and the sample. When the scintillating film is excited by a low-energy electron beam (about 1 KeV), it emits energy that is transferred to the sample, causing the sample to radiate. This luminescence is recorded and correlated to the electron beam position to form an image that is not restricted by the optical diffraction limit.

    Developing the scintillating film and integrating it into a microchip imaging device was an enormous undertaking, Ginsberg says, and she credits the “talent and dedication” of her research group for the success. She also gives much credit to the staff and capabilities of the Molecular Foundry, a DOE Office of Science User Facility, where the CLAIRE imaging demonstration was carried out.

    “The Molecular Foundry truly enabled CLAIRE imaging to come to life,” she says. “We collaborated with staff scientists there to design and install a high efficiency light collection apparatus in one of the Foundry’s scanning electron microscopes and their advice and input were fantastic. That we can work with Foundry scientists to modify the instrumentation and enhance its capabilities not only for our own experiments but also for other users is unique.”

    While there is still more work to do to make CLAIRE widely accessible, Ginsberg and her group are moving forward with further refinements for several specific applications.

    “We’re interested in non-invasively imaging soft functional materials like the active layers in solar cells and light-emitting devices,” she says. “It is especially true in organics and organic/inorganic hybrids that the morphology of these materials is complex and requires nanoscale resolution to correlate morphological features to functions.”

    Ginsberg and her group are also working on the creation of liquid cells for observing biomolecular interactions under physiological conditions. Since electron microscopes can only operate in a high vacuum, as molecules in the air disrupt the electron beam, and since liquids evaporate in high vacuum, aqueous samples must either be freeze-dried or hermetically sealed in special cells.

    “We need liquid cells for CLAIRE to study the dynamic organization of light-harvesting proteins in photosynthetic membranes,” Ginsberg says. “We should also be able to perform other studies in membrane biophysics to see how molecules diffuse in complex environments, and we’d like to be able to study molecular recognition at the single molecule level.”

    In addition, Ginsberg and her group will be using CLAIRE to study the dynamics of nanoscale systems for soft materials in general.

    “We would love to be able to observe crystallization processes or to watch a material made of nanoscale components anneal or undergo a phase transition,” she says. “We would also love to be able to watch the electric double layer at a charged surface as it evolves, as this phenomenon is crucial to battery science.”

    A paper describing the most recent work on CLAIRE has been published in the journal Nano Letters. The paper is titled Cathodoluminescence-Activated Nanoimaging: Noninvasive Near-Field Optical Microscopy in an Electron Microscope. Ginsberg is the corresponding author. Other authors are Connor Bischak, Craig Hetherington, Zhe Wang, Jake Precht, David Kaz and Darrell Schlom.

    This research was primarily supported by the DOE Office of Science and by the National Science Foundation.

    See the full article here.

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition

    A U.S. Department of Energy National Laboratory Operated by the University of California

    University of California Seal

    DOE Seal

  • richardmitnick 12:18 pm on March 28, 2015 Permalink | Reply
    Tags: , Electron Microscopy,   

    From Scripps: “Team Breaks Imaging Barrier” 


    Scripps Research Institute

    March 30, 2015
    Madeline McCurry-Schmidt

    Advances in Electron Microscopy Could Aid Drug Design

    A team from the Carragher lab has imaged a protein complex at the highest resolution ever achieved with single particle cryo-electron microscopy. The image reveals individual molecules at 2.8 Å and is, to the researchers’ knowledge, the first published research using this technique that shows individual water molecules.

    Scientists at The Scripps Research Institute (TSRI) have broken a major barrier in structural imaging. Their study, published recently in the journal eLife, shows a protein complex at the highest resolution ever achieved with a standard technique called single particle cryo-electron microscopy.

    “The instruments and software are now so good that we do not know what the barriers are any more,” said Bridget Carragher, a professor at TSRI with a joint appointment at the New York Structural Biology Center.

    With single particle cryo-electron microscopy, scientists freeze a sample and then expose it to a beam of high-energy electrons. This excites electrons in the sample, allowing scientists to capture an image.

    While the technique has many practical advantages over other structural biology methods, scientists have so far not been able to reach resolutions more detailed than 3 Angstroms (one ten-billionth of a meter, marked with the symbol Å). At this resolution, some of the details of the structure that are important for guiding drug design are not discernable.

    The new study shows that reaching resolutions greater than 3 Å is possible using single particle cryo-electron microscopy. The imaged protein complex reveals individual molecules at 2.8 Å and is, to the researchers’ knowledge, the first time a paper has been published showing individual water molecules using this technique.

    Better Imaging, Better Drugs

    The scientists used a new type of electron microscope, called the FEI Titan Krios, and a new-generation camera, called a Gatan K2 Summit, to break the 3 Å barrier.

    Titan Krios

    Gatan K2 Summit

    The FEI Titan Krios is housed on TSRI’s La Jolla, California, campus. It has a higher energy electron source and a more stable platform than other types of electron microscopes. It also operates with software developed at TSRI through the National Resource for Automated Molecular Microscopy to find the best parts of a sample for imaging.

    The Gatan K2 Summit camera improves imaging by directly detecting electrons, instead of losing resolution by converting electrons to light. The camera can also capture a series of images, essentially a movie, giving scientists the ability to correct for movements in the specimen and make the images as sharp as possible.

    Revealing high-resolution details in a structure helps researchers develop new drugs to treat disease. Structures seen at greater than 3 Å might show vulnerabilities in a virus where drugs could bind, for example.

    “By seeing everything in more detail, you can design more effective drugs,” said Melody Campbell, a TSRI graduate student and co-first author of the new paper with David Veesler, previously a post-doctoral fellow at TSRI and now an assistant professor at the University of Washington.

    The advances in single particle cryo-electron microscopy also allow scientists to image more kinds of structures, more quickly. For many years, scientists have relied on a high-resolution imaging technique called X-ray crystallography. Although X-ray crystallography has led to many advances in drug design, figuring out how to grow a crystal can take years and not all structures can be crystallized.

    Electron microscopy does not require a crystal, however, and many projects take only weeks or months.

    In the new study, the researchers imaged a protein complex from a microbe called Thermoplasma acidophilum. This protein complex, called a proteasome, is also found in humans and is an important target for treating many types of cancer.

    The team spent several months setting up the instruments—since the FEI Titan Krios was new to the institute—and then they captured all the raw data over a single weekend. They then used computational programs to select the clearest images and refine them over several months to build a 3D model of the proteasome.

    “It was a relief to know we had finally done it,” said Campbell. “Now we hope other people can just hop on the microscope, use similar strategies and also get high-resolution structures.”

    In addition to Carragher, Campbell and Veesler, authors of the study, “2.8 Å resolution reconstruction of the Thermoplasma acidophilum 20 S proteasome using cryo-electron microscopy,” were Anchi Cheng and Clinton S. Potter of the New York Structural Biology Center. For more information on the paper, see http://elifesciences.org/content/4/e06380.

    This research was supported by the National Institutes of Health’s National Institute of General Medical Sciences (grant GM103310), a FP7 Marie Curie IOF fellowship (273427) and an American Heart Association fellowship (14PRE18870036).

    See the full article here.

    Please help promote STEM in your local schools.

    STEM Icon

    Stem Education Coalition

    The Scripps Research Institute (TSRI), one of the world’s largest, private, non-profit research organizations, stands at the forefront of basic biomedical science, a vital segment of medical research that seeks to comprehend the most fundamental processes of life. Over the last decades, the institute has established a lengthy track record of major contributions to the betterment of health and the human condition.

    The institute — which is located on campuses in La Jolla, California, and Jupiter, Florida — has become internationally recognized for its research into immunology, molecular and cellular biology, chemistry, neurosciences, autoimmune diseases, cardiovascular diseases, virology, and synthetic vaccine development. Particularly significant is the institute’s study of the basic structure and design of biological molecules; in this arena TSRI is among a handful of the world’s leading centers.

    The institute’s educational programs are also first rate. TSRI’s Graduate Program is consistently ranked among the best in the nation in its fields of biology and chemistry.

  • richardmitnick 3:31 pm on January 28, 2013 Permalink | Reply
    Tags: , , Electron Microscopy,   

    From PNNL Lab: “Seeing a Common Catalyst with New Eyes” 

    Chemical imaging microscope shows corrugated gamma-alumina surface
    January 2013
    Suraiya Farukhi
    Christine Sharp

    Results: Neither smooth nor disordered, gamma-alumina nanoparticles are corrugated with tiny pores inside, according to scientists at Pacific Northwest National Laboratory. Using a powerful transmission electron microscope, the team obtained ultrahigh-resolution images and chemical data about the particle’s surface. They found that the particles were covered with ridges made from a more open, yet symmetrical, arrangement of atoms. The open arrangement on the surfaces, notated as (110), covers 70% of the nanoparticle.

    The surface of the plate-like particles is far from smooth, according to a new transmission electron microscopy study conducted by Pacific Northwest National Laboratory and the FEI Company.

    By understanding the structure and function of tiny gamma-alumina particles, scientists are taking crucial steps to optimizing and realizing new useful properties for these materials. ‘If we can learn about the surfaces, then we can tailor them and make them more efficient in catalytic applications,’ said Dr. Libor Kovarik, who led the imaging study as part of PNNL’s Chemical Imaging Initiative.

    Why It Matters: Reducing refineries’ energy demands or car and truck emissions requires efficient catalysts on durable support materials. The supporting material must withstand severe temperature and pressure changes. Gamma-alumina has been studied extensively, but its atomic arrangement has not been established because of the challenge of getting a detailed view of this complex material. Accurately describing the atomic structure is crucial for understanding and taking advantage of the best properties of gamma-alumina.

    ‘Catalytic research demands this type of state-of-the-art chemical imaging research,’ said Dr. Charles Peden, a heterogeneous catalysis scientist who worked on the study, and an Associate Director of PNNL’s Institute for Integrated Catalysis. ‘Dr. Kovarik’s outstanding new images from this powerhouse microscope have yielded unprecedented new information about a catalyst material of enormous practical utility.'”

    See the full article here.

    Pacific Northwest National Laboratory (PNNL) is one of the United States Department of Energy National Laboratories, managed by the Department of Energy’s Office of Science. The main campus of the laboratory is in Richland, Washington.

    PNNL scientists conduct basic and applied research and development to strengthen U.S. scientific foundations for fundamental research and innovation; prevent and counter acts of terrorism through applied research in information analysis, cyber security, and the nonproliferation of weapons of mass destruction; increase the U.S. energy capacity and reduce dependence on imported oil; and reduce the effects of human activity on the environment. PNNL has been operated by Battelle Memorial Institute since 1965.


    ScienceSprings is powered by MAINGEAR computers

  • richardmitnick 1:14 pm on January 17, 2013 Permalink | Reply
    Tags: , , Electron Microscopy   

    From Berkeley Lab: “New Key to Organism Complexity Identified” 

    Berkeley Lab

    Berkeley Scientists Find that a Critical Transcription Factor Co-exists in Two Distinct States

    January 17, 2013
    Lynn Yarris

    The enormously diverse complexity seen amongst individual species within the animal kingdom evolved from a surprisingly small gene pool. For example, mice effectively serve as medical research models because humans and mice share 80-percent of the same protein-coding genes.

    The ‘rearranged’ state of the lobe A (yellow) of the horseshoe-like TFIID transcription factor enables TFIID to bind with DNA (green) and start the process by which DNA is copied into RNA.

    The key to morphological and behavioral complexity, a growing body of scientific evidence suggests, is the regulation of gene expression by a family of DNA-binding proteins called ‘transcription factors.’ Now, a team of researchers with the U.S. Department of Energy’s Lawrence Berkeley National Laboratory (Berkeley Lab) and the University of California (UC) Berkeley has discovered the secret behind how one these critical transcription factors is able to perform – a split personality.

    Using a technique called single-particle cryo-electron microscopy, the team, which was led by biophysicist Eva Nogales, showed that the transcription factor known as ‘TFIID’ can co-exist in two distinct structural states.”

    two people
    Michael Cianfrocco and Eva Nogales used single-particle cryo-electron microscopy to learn how the TFIID transcription factor helps regulate of gene expression, a process critical to the growth, development, health and survival of all organisms. (Photo by Roy Kaltschmidt)

    See the full article here.

    A U.S. Department of Energy National Laboratory Operated by the University of California


    ScienceSprings is powered by MAINGEAR computers

Compose new post
Next post/Next comment
Previous post/Previous comment
Show/Hide comments
Go to top
Go to login
Show/Hide help
shift + esc
%d bloggers like this: