Tristan Al-Haddad, left, assistant professor in the Georgia Tech College of Architecture, and College of Architecture research scientist Karl Brohammer, examine architectural cladding prototypes that have been digitally fabricated using ultra-high-performance concrete. (Click image for high-resolution version. Credit: Gary Meek)

Researchers in the College of Architecture at the Georgia Institute of Technology are now automating some of the processes by which computer-based designs are turned into real world entities. They’re developing techniques that fabricate building elements directly from digital designs, allowing custom concrete components to be manufactured rapidly and at low cost.

“We’re developing the research and the protocols to manufacture high-end customized architectural products economically, safely and with environmental responsibility,” said Tristan Al-Haddad, an assistant professor in the College of Architecture who is a leader in this effort. “We think this work offers opportunities for architectural creativity at a new level and with tremendously increased efficiency.”

• In one recent project, Al-Haddad and a College of Architecture team collaborated with Lafarge North America to fabricate an award-winning building-element concept called a “Liquid Wall.” The Georgia Tech team employed digital techniques to help construct a prototype wall, using ultra high-performance concrete; the result was displayed by the New York Chapter of the American Institute of Architects (AIANY) in the “Innovate:Integrate” exhibition.
• In another Lafarge-sponsored project, Al-Haddad and a College of Architecture team are developing a complete free-standing structure using ultra high-performance concrete elements fabricated directly from digital designs.

Computing a Wall

The Liquid Wall, originated by Peter Arbour of Paris-based RFR Consulting Engineers, won the 2010 Open Call for Innovative Curtain-Wall Design competition conducted by the AIA. The concept advanced a novel approach to curtain walls, which are building coverings that keep out weather but are non-structural and lightweight.

Researchers in Georgia Tech’s College of Architecture are helping automate the process of turning CAD designs into manufactured products. Here, professor Tristan Al-Haddad and undergraduate students Sam Kim and Patrick di Rito are evaluating custom wall structures manufactured using a new process. (Click image for high-resolution version. Credit: Gary Meek).

RFR’s plans called for the Liquid Wall to be constructed of stainless steel and Ductal®, a light and strong ultra-high-performance concrete (UHPC) that is produced by Lafarge. Moreover, the new building enclosure was conceived as an entire system, including integrated louver systems, solar shading, integrated passive solar collectors and other advanced features.

Georgia Tech became involved in the Liquid Wall project when RFR decided to built a full-scale prototype of the complex concept. RFR asked Al-Haddad to help turn Arbour’s original parametric sketches into a manufacturable design.

Supported by the College of Architecture’s Digital Building and Digital Fabrication laboratories, the researchers refined the geometry of the original sketches for manufacturability and developed the techniques required for fabricating a full-size curtain wall. Then, working from their digital models and using a five-axis CNC router – a device capable of machining material directly from a digital design – the Georgia Tech team milled a full-scale model of the wall. The model was made from a lightweight polymer material, expanded polystyrene (EPS) closed-cell foam, which was then given a polyurea coating.

The digitally milled foam model created an exact replica — a positive — of the final wall.  The lightweight positive could then be used to produce a negative capable of forming the actual prototype. In this case, the collaborators used the positive to produce a rubber mold – the negative – from which the final wall was cast.

The foam positive was shipped to Coreslab Structures Inc., a large corporation that specializes in industrial-scale casting. The Georgia Tech team then worked with Coreslab to identify the best techniques for creating the rubber mold and for pouring in Ductal to form the concrete wall.

“It was a very collaborative process – the four major players were Peter Arbour and RFR, Georgia Tech, Coreslab and Lafarge,” Al-Haddad said. “And we had all of three weeks to finish the work before the exhibition deadline – so it was pretty intense.”

Other College of Architecture people involved in the collaboration included graduate student Andres Cavieres, associate professor Russell Gentry and professor Charles Eastman, director of the Digital Building Laboratory. The resulting full-size Liquid Wall prototype was installed at the Center for Architecture in New York City as part of the AIANY’s “Innovate: Integrate” exhibition, and was on view for several months in 2010 and 2011.

Confronting Challenges

The Liquid Wall project was challenging, said Eastman, who holds joint appointments in the College of Architecture and the College of Computing. The process involved not only producing rubber negatives using wall-form designs created with CAD and parametric-modeling software, but also required identifying the right production procedures and finding effective ways of installing a completed full-size wall on a building.

“When you’re creating a completely new process like the Liquid Wall, you’re faced with developing a whole new manufacturing process for this kind of material,” Eastman said.

A future project, expected to be about 20 by 20 feet square and 15 feet high, will be built using Ductal UHPC, principally or entirely. A central technical challenge will involve molding the many custom elements so that all edges fit together and form a structure that is stable, practical and esthetically pleasing.

“We understand the structural side of a project like this quite well — the difficulty comes in the actual manufacturing of the elements,” Al-Haddad said. “We want to advance the use of digital parametric models with custom molding systems, and create a free-form manufacturing system that can produce many variations quickly and accurately.”

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Hematoxylin and eosin (H&E) staining of sections of wild-type (top row) and H1 triple-knockout (bottom row) embryoid bodies. After 14 days in rotary suspension culture, the wild-type embryoid bodies showed more differentiated morphologies with cysts forming (black arrows) and the knockout embryoid bodies failed to form cavities (far right). (Click image for high-resolution version. Credit: Yuhong Fan)

Researchers from the Georgia Institute of Technology and Emory University found that chromatin compaction is required for proper embryonic stem cell differentiation to occur. Chromatin, which is composed of histone proteins and DNA, packages DNA into a smaller volume so that it fits inside a cell.

A study published on May 10, 2012 in the journal PLoS Genetics found that embryonic stem cells lacking several histone H1 subtypes and exhibiting reduced chromatin compaction suffered from impaired differentiation under multiple scenarios and demonstrated inefficiency in silencing genes that must be suppressed to induce differentiation.

“While researchers have observed that embryonic stem cells exhibit a relaxed, open chromatin structure and differentiated cells exhibit a compact chromatin structure, our study is the first to show that this compaction is not a mere consequence of the differentiation process but is instead a necessity for differentiation to proceed normally,” said Yuhong Fan, an assistant professor in the Georgia Tech School of Biology.

Fan and Todd McDevitt, an associate professor in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University, led the study with assistance from Georgia Tech graduate students Yunzhe Zhang and Kaixiang Cao, research technician Marissa Cooke, and postdoctoral fellow Shiraj Panjwani.

The work was supported by the National Institutes of Health’s National Institute of General Medical Sciences (NIGMS), the National Science Foundation, a Georgia Cancer Coalition Distinguished Scholar Award, and a Johnson & Johnson/Georgia Tech Healthcare Innovation Award.

Phase contrast images showing that H1 triple-knockout (bottom) embryonic stem cells were unable to adequately form neurites and neural networks compared to wild-type embryonic stem cells (top). (Click image for high-resolution version. Credit: Yuhong Fan)

To investigate the impact of linker histones and chromatin folding on stem cell differentiation, the researchers used embryonic stem cells that lacked three subtypes of linker histone H1 — H1c, H1d and H1e — which is the structural protein that facilitates the folding of chromatin into a higher-order structure. They found that the expression levels of these H1 subtypes increased during embryonic stem cell differentiation, and embryonic stem cells lacking these H1s resisted spontaneous differentiation for a prolonged time, showed impairment during embryoid body differentiation and were unsuccessful in forming a high-quality network of neural cells.

“This study has uncovered a new, regulatory function for histone H1, a protein known mostly for its role as a structural component of chromosomes,” said Anthony Carter, who oversees epigenetics grants at NIGMS.  “By showing that H1 plays a part in controlling genes that direct embryonic stem cell differentiation, the study expands our understanding of H1’s function and offers valuable new insights into the cellular processes that induce stem cells to change into specific cell types.”

During spontaneous differentiation, the majority of the H1 triple-knockout embryonic stem cells studied by the researchers retained a tightly packed colony structure typical of undifferentiated cells and expressed high levels of Oct4 for a prolonged time. Oct4 is a pluripotency gene that maintains an embryonic stem cell’s ability to self-renew and must be suppressed to induce differentiation.

“H1 depletion impaired the suppression of the Oct4 and Nanog pluripotency genes, suggesting a novel mechanistic link by which H1 and chromatin compaction may mediate pluripotent stem cell differentiation by contributing to the epigenetic silencing of pluripotency genes,” explained Fan. “While a significant reduction in H1 levels does not interfere with embryonic stem cell self-renewal, it appears to impair differentiation.”

Immunostaining of wild-type (top) and H1 triple-knockout (bottom) cultures under a neural differentiation protocol. The H1 triple-knockout cells were defective in forming neuronal and glial cells and a neural network, which is essential for nervous system development. (Click image for high-resolution version. Credit: Yuhong Fan)

The researchers also used a rotary suspension culture method developed by McDevitt to produce with high efficiency homogonous 3D clumps of embryonic stem cells called embryoid bodies. Embryoid bodies typically contain cell types from all three germ layers — the ectoderm, mesoderm and endoderm — that give rise to the various types of tissues and structures in the body. However, the majority of the H1 triple-knockout embryoid bodies formed in rotary suspension culture lacked differentiated structures and displayed gene expression signatures characteristic of undifferentiated stem cells.

“H1 triple-knockout embryoid bodies displayed a reduced level of activation of many developmental genes and markers in rotary culture, suggesting that differentiation to all three germ layers was affected.” noted McDevitt.

The embryoid bodies also lacked the epigentic changes at the pluripotency genes necessary for differentiation, according to Fan.

“When we added one of the deleted H1 subtypes to the embryoid bodies, Oct4 was suppressed normally and embryoid body differentiation continued,” explained Fan. “The epigenetic regulation of Oct4 expression by H1 was also evident in mouse embryos.”

In another experiment, the researchers provided an environment that would encourage embryonic stem cells to differentiate into neural cells. However, the H1 triple-knockout cells were defective in forming neuronal and glial cells and a neural network, which is essential for nervous system development. Only 10 percent of the H1 triple-knockout embryoid bodies formed neurites and they produced on average eight neurites each. In contrast, half of the normal embryoid bodies produced, on average, 18 neurites.

In future work, the researchers plan to investigate whether controlling H1 histone levels can be used to influence the reprogramming of adult cells to obtain induced pluripotent stem cells, which are capable of differentiating into tissues in a way similar to embryonic stem cells.

Research reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health (NIH) under award number GM085261 and the National Science Foundation under award number CBET-0939511. The content is solely the responsibility of the principal investigators and does not necessarily represent the official views of the NIH or NSF.

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Jonathan Colton, a professor in the George W. Woodruff School of Mechanical Engineering and the School of Industrial Design at Georgia Tech, will pursue an innovative global health and development research project focused on designing a net-zero energy warehousing and distribution system for vaccines and drugs in developing countries. Net-zero energy describes a building with no net energy consumption and no carbon emissions measured on an annual basis.

Jonathan Colton, a professor in the School of Mechanical Engineering and the School of Industrial Design at Georgia Tech, received a $100,000 Grand Challenges Explorations grant from the Bill & Melinda Gates Foundation to design a net-zero energy warehousing and distribution system for vaccines and drugs in developing countries. (Click image for high-resolution version. Credit: USAID/PATH/Gabe Bienczycki)

In addition to Colton, immunization logistics consultant John Lloyd, architect Andrew Garnett and Solar Electric Light Fund project manager Steve McCarney will also contribute to the project.

The project was one of more than 100 Grand Challenges Explorations grants announced May 9, 2012.

“Grand Challenges Explorations encourages individuals worldwide to expand the pipeline of ideas where creative, unorthodox thinking is most urgently needed,” said Chris Wilson, director of Global Health Discovery and Translational Sciences at the Bill & Melinda Gates Foundation.  “We’re excited to provide additional funding for select grantees so that they can continue to advance their idea towards global impact.”

The goal of the Georgia Tech project is to develop the design and engineering specifications for a new, energy-optimized warehousing and distribution system for vaccines and drugs. In low- and middle-income countries, vaccines and drugs are often stored in older buildings that are inefficiently laid out and wasteful of energy. In these countries, warehousing and distribution costs can amount to 20 percent of drug and vaccine supply costs.

“We plan to demonstrate that energy-efficient, state-of-the-art warehousing systems can eliminate or greatly reduce the operational energy costs for storage and distribution of vaccines and drugs in developing countries with challenging climates,” said Colton.

According to Colton, to be successful the new warehousing system will need to:

• Minimize environmental impact, energy consumption, and storage and transport costs;
• Offset any grid electricity consumption;
• Employ low-energy cooling techniques;
• Accommodate a variety of building sizes and configurations; and
• Be able to store vaccines, drugs and dry supplies at various controlled temperatures.

“Once we create the design and engineering specifications for this new warehousing and storage system, we plan to select an industry partner to build and test the system in a developing country such as Tunisia,” added Colton.

About Grand Challenges Explorations: Grand Challenges Explorations is a $100 million initiative funded by the Bill & Melinda Gates Foundation. Launched in 2008, more than 600 people in 45 countries have received Grand Challenges Explorations grants. The grant program is open to anyone from any discipline and from any organization. The initiative uses an agile, accelerated grant-making process with short two-page online applications and no preliminary data required. Initial grants of $100,000 are awarded two times a year. Successful projects have the opportunity to receive a follow-on grant of up to $1 million.

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Researchers at MIT and Georgia Tech have developed a way to automate a process called whole-cell patch clamping, which involves bringing a tiny hollow glass pipette in contact with the cell membrane of a neuron, then opening up a small pore in the membrane to record the electrical activity within the cell. (Click image for high-resolution version. Credit: Sputnik Animation and MIT McGovern Institute)

But that could soon change: Researchers at MIT and the Georgia Institute of Technology have developed a way to automate the process of finding and recording information from neurons in the living brain. The researchers have shown that a robotic arm guided by a cell-detecting computer algorithm can identify and record from neurons in the living mouse brain with better accuracy and speed than a human experimenter.

The new automated process eliminates the need for months of training and provides long-sought information about living cells’ activities. Using this technique, scientists could classify the thousands of different types of cells in the brain, map how they connect to each other, and figure out how diseased cells differ from normal cells.

The project is a collaboration between the labs of Ed Boyden, associate professor of biological engineering and brain and cognitive sciences at MIT, and Craig Forest, an assistant professor in the George W. Woodruff School of Mechanical Engineering at Georgia Tech.

“Our team has been interdisciplinary from the beginning, and this has enabled us to bring the principles of precision machine design to bear upon the study of the living brain,” Forest says. His graduate student, Suhasa Kodandaramaiah, spent the past two years as a visiting student at MIT, and is the lead author of the study, which appears in the May 6 issue of Nature Methods.

The method could be particularly useful in studying brain disorders such as schizophrenia, Parkinson’s disease, autism and epilepsy, Boyden says. “In all these cases, a molecular description of a cell that is integrated with [its] electrical and circuit properties … has remained elusive,” says Boyden, who is a member of MIT’s Media Lab and McGovern Institute for Brain Research. “If we could really describe how diseases change molecules in specific cells within the living brain, it might enable better drug targets to be found.”

Automation

Kodandaramaiah, Boyden and Forest set out to automate a 30-year-old technique known as whole-cell patch clamping, which involves bringing a tiny hollow glass pipette in contact with the cell membrane of a neuron, then opening up a small pore in the membrane to record the electrical activity within the cell. This skill usually takes a graduate student or postdoc several months to learn.

MIT and Georgia Tech researchers developed a four-step process that a robotic arm guided by a cell-detecting computer algorithm uses to find and record information from neurons in the living brain. The pipette is lowered to a target zone in the brain, the pipette is advanced until a neuron is detected, a seal is formed between the pipette and the cell, and a small pore is opened in the membrane to record the electrical activity within the cell. (Click image for high-resolution version. Credit: MIT and Georgia Tech)

Kodandaramaiah spent about four months learning the manual patch-clamp technique, giving him an appreciation for its difficulty. “When I got reasonably good at it, I could sense that even though it is an art form, it can be reduced to a set of stereotyped tasks and decisions that could be executed by a robot,” he says.

To that end, Kodandaramaiah and his colleagues built a robotic arm that lowers a glass pipette into the brain of an anesthetized mouse with micrometer accuracy. As it moves, the pipette monitors a property called electrical impedance — a measure of how difficult it is for electricity to flow out of the pipette. If there are no cells around, electricity flows and impedance is low. When the tip hits a cell, electricity can’t flow as well and impedance goes up.

The pipette takes two-micrometer steps, measuring impedance 10 times per second. Once it detects a cell, it can stop instantly, preventing it from poking through the membrane. “This is something a robot can do that a human can’t,” Boyden says.

Once the pipette finds a cell, it applies suction to form a seal with the cell’s membrane. Then, the electrode can break through the membrane to record the cell’s internal electrical activity. The robotic system can detect cells with 90 percent accuracy, and establish a connection with the detected cells about 40 percent of the time.

The researchers also showed that their method can be used to determine the shape of the cell by injecting a dye; they are now working on extracting a cell’s contents to read its genetic profile.

Development of the new technology was funded primarily by the National Institutes of Health, the National Science Foundation and the MIT Media Lab.

New era for robotics

The researchers recently created a startup company, Neuromatic Devices, to commercialize the device.

MIT researcher Ed Boyden (left) and Georgia Tech researchers Suhasa Kodandaramaia (seated) and Craig Forest have developed a way to automate the process of finding and recording information from neurons in the living brain. (Click image for high-resolution version. Credit: MIT)

The researchers are now working on scaling up the number of electrodes so they can record from multiple neurons at a time, potentially allowing them to determine how different parts of the brain are connected.

They are also working with collaborators to start classifying the thousands of types of neurons found in the brain. This “parts list” for the brain would identify neurons not only by their shape — which is the most common means of classification — but also by their electrical activity and genetic profile.

“If you really want to know what a neuron is, you can look at the shape, and you can look at how it fires. Then, if you pull out the genetic information, you can really know what’s going on,” Forest says. “Now you know everything. That’s the whole picture.”

Boyden says he believes this is just the beginning of using robotics in neuroscience to study living animals. A robot like this could potentially be used to infuse drugs at targeted points in the brain, or to deliver gene therapy vectors. He hopes it will also inspire neuroscientists to pursue other kinds of robotic automation — such as in optogenetics, the use of light to perturb targeted neural circuits and determine the causal role that neurons play in brain functions.

Neuroscience is one of the few areas of biology in which robots have yet to make a big impact, Boyden says. “The genome project was done by humans and a giant set of robots that would do all the genome sequencing. In directed evolution or in synthetic biology, robots do a lot of the molecular biology,” he says. “In other parts of biology, robots are essential.”

Other co-authors include MIT grad student Giovanni Talei Franzesi and MIT postdoc Brian Y. Chow.

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Georgia Tech Research Institute researchers Brent Wagner (l) and Bernd Kahn are using novel materials and nanotechnology techniques to develop improved radiation detection. (Click image for high-resolution version. Credit: Gary Meek)

To support the nation’s nuclear-surveillance capabilities, researchers at the Georgia Tech Research Institute (GTRI) are developing ways to enhance the radiation-detection devices used at ports, border crossings, airports and elsewhere. The aim is to create technologies that will increase the effectiveness and reliability of detectors in the field, while also reducing cost. The work is co-sponsored by the Domestic Nuclear Defense Office of the Department of Homeland Security and by the National Science Foundation.

“U.S. security personnel have to be on guard against two types of nuclear attack — true nuclear bombs, and devices that seek to harm people by dispersing radioactive material,” said Bernd Kahn, a researcher who is principal investigator on the project. “Both of these threats can be successfully detected by the right technology.”

The GTRI team, led by co-principal investigator Brent Wagner, is utilizing novel materials and nanotechnology techniques to produce improved radiation detection. The researchers have developed the Nano-photonic Composite Scintillation Detector, a prototype that combines rare-earth elements and other materials at the nanoscale for improved sensitivity, accuracy and robustness.

Bernd Kahn, seated, and Brent Wagner review a pulse height spectrum collected using a scintillation radiation detector they devised. The GTRI researchers are using novel materials and nanotechnology techniques to develop improved radiation detection. (Click image for high-resolution version. Credit: Gary Meek)

Details of the research were presented April 23, 2012 at the SPIE Defense, Security, and Sensing Conference held in Baltimore, MD.

Scintillation detectors and solid-state detectors are two common types of radiation detectors, Wagner explained. A scintillation detector commonly employs a single crystal of sodium iodide or a similar material, while a solid-state detector is based on semiconducting materials such as germanium.

Both technologies are able to detect gamma rays and subatomic particles emitted by nuclear material. When gamma rays or particles strike a scintillation detector, they create light flashes that are converted to electrical pulses to help identify the radiation at hand. In a solid-state detector, incoming gamma rays or particles register directly as electrical pulses.

“Each reaction to a gamma ray takes a very short time — a fraction of a microsecond,” Wagner said. “By looking at the number and the intensity of the pulses, along with other factors, we can make informed judgments about the type of radioactive material we’re dealing with.”

But both approaches have drawbacks. A scintillation detector requires a large crystal grown from sodium iodide or other materials. Such crystals are typically fragile, cumbersome, difficult to produce and extremely vulnerable to humidity.

Examples of scintillators that were produced from molten glass by the GTRI researchers. The wormlike blue structure is an artifact from the glass-molding process. (Click image for high-resolution version. Credit: Gary Meek)

A germanium-based solid-state detector offers better identification of different kinds of nuclear materials. But high-purity single-crystal germanium is difficult to make in a large volume; the result is less-sensitive devices with reduced ability to detect radiation at a distance. Moreover, germanium must be kept extremely cold — 200 degrees below zero Celsius — to function properly, which poses problems for use in the field.

The Nanoscale Advantage

To address these problems, the GTRI team has been investigating a wide variety of alternative materials and methodologies. After selecting the scintillation approach over solid-state, the researchers developed a composite material — composed of nanoparticles of rare-earth elements, halides and oxides — capable of creating light.

“A nanopowder can be much easier to make, because you don’t have to worry about producing a single large crystal that has zero imperfections,” Wagner said.

A scintillator crystal must be transparent to light, he explained, a quality that’s key to its ability to detect radiation. A perfect crystal uniformly converts incoming energy from gamma rays to flashes of light. A photo-multiplier then amplifies these flashes of light so they can be accurately measured to provide information about radioactivity.

Georgia Tech Research Institute researchers Brent Wagner (l) and Bernd Kahn are using novel materials and nanotechnology techniques to develop improved radiation detection. (Click image for high-resolution version. Credit: Gary Meek)

However, when a transparent material — such as crystal or glass — is ground into smaller pieces, its transparency disappears. As a result, a mixture of particles in a transparent glass would scatter the luminescence created by incoming gamma rays. That scattered light can’t reach the photo-multiplier in a uniform manner, and the resulting readings are badly skewed.

To overcome this issue, the GTRI team reduced the particles to the nanoscale. When a nanopowder reaches particle sizes of 20 nanometers or less, scattering effects fade because the particles are now significantly smaller than the wavelength of incoming gamma rays.

“Think of it as a big ocean wave coming in,” Wagner said. “That wave would definitely interact with a large boat, but something the size of a beach ball doesn’t affect it.”

Rare Earths and Silica

At first the team worked on dispersing radiation-sensitive crystalline nanoparticles in a plastic matrix. But they encountered problems with distributing the nanopowder uniformly enough in the matrix to achieve sufficiently accurate radiation readings.

More recently, the researchers have investigated a parallel path using glass rather than plastic as a matrix material, combining gadolinium and cerium bromide with silica and alumina.

Kahn explained that gadolinium or a similar material is essential to scintillation-type particle detection because of its role as an absorber. But in this case, when an incoming gamma ray is absorbed in gadolinium, the energy is not efficiently emitted in the form of luminescence.

Instead, the light emission role here falls to a second component — cerium. The gadolinium absorbs energy from an incoming gamma ray and transfers that energy to the cerium atom, which then acts as an efficient light emitter.

The researchers found that by heating gadolinium, cerium, silica and alumina and then cooling them from a molten mix to a solid monolith, they could successfully distribute the gadolinium and cerium in silica-based glasses. As the material cools, gadolinium and cerium precipitate out of the aluminosilicate solution and are distributed throughout the glass in a uniform manner. The resulting composite gives dependable readings when exposed to incoming gamma rays.

“We’re optimistic that we’ve identified a productive methodology for creating a material that could be effective in the field,” Wagner said. “We’re continuing to work on issues involving purity, uniformity and scaling, with the aim of producing a material that can be successfully tested and deployed.”

This material is based upon work supported by the U.S. Department of Homeland Security under Grant Award Number 2008-DN-077-ARI001-02. The views and conclusions contained in this document are those of the authors and should not be interpreted as necessarily representing the official policies, either expressed or implied, of the U.S. Department of Homeland Security.

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Molecular probes displaying the LNLPHG and RFSAFY peptide sequences showed the greatest binding affinity to fibronectin fibers and the greatest efficiency in discriminating between relaxed and strained fibers. On extracellular matrix assembled by primary lung fibroblasts, LNLPHG preferentially attached to relaxed fibronectin fibers (top row), whereas RFSAFY bound to strained fibers (bottom row). (Scale bar: 20 microns) (Click image for high-resolution version. Credit: Thomas Barker)

During physiological processes, fibronectin fibers are believed to experience mechanical forces that strain the fibers and cause dramatic structural modifications that change their biological activity. While understanding the role of fibronectin strain events in development and disease progression is becoming increasingly important, detecting and interrogating these events is difficult.

In a new study, researchers identified molecular probes capable of selectively attaching to fibronectin fibers under different strain states, enabling the detection and examination of fibronectin strain events in both culture and living tissues.

“The mechano-sensitive molecular probes we identified allow us to dynamically examine the relevance of mechanical strain events within the natural cellular microenvironment and correlate these events with specific alterations in fibronectin associated with the progression of disease,” said Thomas Barker, an assistant professor in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University.

The study was published on April 23, 2012 in the online early edition of the journal Proceedings of the National Academy of Sciences. Barker worked on the study with Georgia Tech graduate student Lizhi Cao and Harry Bermudez, an assistant professor in the University of Massachusetts Amherst Department of Polymer Science and Engineering. The research was supported by the National Institutes of Health.

Researchers have hypothesized that mechanical forces emanating from cells may partially unfold fibronectin and regulate what proteins bind to it. While simulation and tissue culture experiments support this hypothesis, direct evidence that such molecular events occur in living organisms has not yet been presented, according to Barker.

A technique called intramolecular fluorescence resonance energy transfer (FRET) has been used to detect molecular strain events in fibronectin fibers, but the technique has limitations because it cannot be used on living tissues and requires the fibronectin to be chemically labeled.

“The molecular probes we identified can be used to map molecular strain events in native extracellular matrix and living lung tissues,” explained Barker. “The probes can also be used to study the mechanism by which cells control the mechanical forces that alter fibronectin’s conformation, control the exposure of its binding sites and regulate cell signaling.”

Staining of fibronectin-targeting molecular probes displaying the LNLPHG (blue) and RFSAFY (red) peptide sequences on prepared living lung slices. The probes did not attach to the same fiber, which confirmed their ability to selectively discriminate strained or relaxed regions within a fibronectin fiber network. (Click image for high-resolution version. Credit: Thomas Barker)

The researchers used a controlled fibronectin fiber deposition and extension technique to apply tension to the fibers and stretch them to 2.6 times their original length without significant breakage. Then they used a technique called phage display to identify peptides capable of discriminating fibronectin fibers under relaxed and strained conditions. The molecular probes displaying peptide sequences LNLPHG and RFSAFY showed the greatest binding affinity to fibronectin fibers and the greatest efficiency in discriminating between relaxed and strained fibers.

For proof-of-concept demonstrations, the researchers used the probes to discriminate fibronectin fibers within native extracellular matrix and mouse lung slices. LNLPHG preferentially attached to relaxed fibronectin fibers, whereas RFSAFY bound to strained fibers. The probes never attached to the same fiber, which confirmed their ability to selectively discriminate regions within a fibronectin fiber network.

“This study strongly suggests that fibronectin fibers under strain display markedly different biochemical signatures that can be used for the molecular-level detection of fibronectin fiber strain,” explained Barker. “The data also show the potential for living tissue to be interrogated for mechano-chemical alterations that lead to physiological and pathological progression.”

In the future, the researchers hope to use these fibronectin strain-sensitive probes to target therapeutics to fibronectin fibers based on their mechanical signature.

This work was supported in part by training grants from the National Institutes of Health (NIH) (Award Nos. T32-GM008433 and T32-EB006343). The content is solely the responsibility of the principal investigators and does not necessarily represent the official views of the NIH.

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Image shows single-molecule identification. The green cross signs show the locations of single molecules using the super resolution technique. (Click image for high-resolution version. Image Courtesy of Lei Zhu and Bo Huang).

Despite many achievements in the field of super-resolution microscopy in the past few years with spatial resolution advances, live-cell imaging has remained a challenge because of the need for high temporal resolution.

Now, Lei Zhu, assistant professor in Georgia Tech’s George W. Woodruff School of Mechanical Engineering, and Bo Huang, assistant professor in UCSF’s Department of Pharmaceutical Chemistry and Department of Biochemistry and Biophysics, have developed an advanced approach using super-resolution microscopy to resolve cellular features an order of magnitude smaller than what could be seen before. This allows the researchers to tap previously inaccessible information and answer new biological questions.

The research was published April 22, 2012 in the journal Nature Methods. The research is funded by the National Institutes of Health, UCSF Program for Breakthrough Biomedical Research, Searle Scholarship and Packard Fellowship for Science and Engineering.

The previous technology using the single-molecule-switching approach for super-resolution microscopy depends on spreading single molecule images sparsely into many, often thousands of, camera frames. It is extremely limited in its temporal resolution and does not provide the ability to follow dynamic processes in live cells.

“We can now use our discovery using super-resolution microscopy with seconds or even sub-second temporal resolution for a large field of view to follow many more dynamic cellular processes,” said Zhu. “Much of our knowledge of the life of a cell comes from our ability to see the small structures within it.”

Huang noted, “One application, for example, is to investigate how mitochondria, the power house of the cell, interact with other organelles and the cytoskeleton to reshape the structure during the life cycle of the cell.”

Currently, light microscopy, especially in the modern form of fluorescence microscopy, is still used frequently by many biologists. However, the authors say, conventional light microscopy has one major limitation: the inability to resolve two objects closer than half the wavelength of the light because of the phenomenon called diffraction. With diffraction, the images look blurry and overlapped no matter how high the magnification that is used.

“The diffraction limit has long been regarded as one of the fundamental constraints for light microscopy until the recent inventions of super-resolution fluorescence microscopy techniques,” said Zhu. Super-resolution microscopy methods, such as stochastic optical reconstruction microscopy (STORM) or photoactivated localization microscopy (PALM), rely on the ability to record light emission from a single molecule in the sample.

Using probe molecules that can be switched between a visible and an invisible state, STORM/PALM determines the position of each molecule of interest. These positions ultimately define a structure.

The new finding is significant, said Zhu and Huang, because they have shown that the technology allows for following the dynamics of a microtubule cytoskeleton with a three-second time resolution, which would allow researchers to study the active transports of vesicles and other cargos inside the cell.

Using the same optical system and detector as in conventional light microscopy, super-resolution microscopy naturally requires longer acquisition time to obtain more spatial information, leading to a trade-off between its spatial and temporal resolution. In super-resolution microscopy methods based on STORM/PALM, each camera image samples a very sparse subset of probe molecules in the sample.

An alternative approach is to increase the density of activated fluorophores so that each camera frame samples more molecules. However, this high density of fluorescent spots causes them to overlap, invalidating the widely used single-molecule localization method.

The authors said that a number of methods have been reported recently that can efficiently retrieve single-molecule positions even when the single fluorophore signals overlap. These methods are based on fitting clusters of overlapped spots with a variable number of point-spread functions (PSFs) with either maximum likelihood estimation or Bayesian statistics. The Bayesian method has also been applied to the whole image set.

As a result of new research, Zhu and Huang present a new approach based on global optimization using compressed sensing, which does not involve estimating or assuming the number of molecules in the image. They show that compressed sensing can work with much higher molecule densities compared to other technologies and demonstrate live cell imaging of fluorescent protein-labeled microtubules with three-second temporal resolution.

The STORM experiment used by the authors, with immunostained microtubules in Drosophila melanogaster S2 cells, demonstrated that nearby microtubules can be resolved by compressed sensing using as few as 100 camera frames, whereas they were not discernible by the single-molecule fitting method. They have also performed live STORM on S2 cells stably expressing tubulin fused to mEos2.

At the commonly used camera frame rate of 56.4 Hertz, a super-resolution movie was constructed with a time resolution of three seconds (169 frames) and a Nyquist resolution of 60 nanometers, much faster than previously reported, said Zhu and Huang. These results have proven that compressed sensing can enable STORM to monitor live cellular processes with second-scale time resolution, or even sub-second-scale resolution if a faster camera can be used.

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Writer: Sarah E. Goodwin

Georgia Tech graduate student Yaroslav Dudin and professor Alex Kuzmich (l-r) adjust optics as part of research into the production of single photons for use in optical quantum information processing and the study of certain physical systems. (Click image for high-resolution version. Credit: John Toon)

The technique takes advantage of the unique properties of atoms that have one or more electrons excited to a condition of near-ionization known as the Rydberg state. Atoms in this highly excited state – with a principal quantum number greater than 70 – have exaggerated electromagnetic properties and interact strongly with one another. That allows one Rydberg atom to block the formation of additional excited atoms within an area of 10 to 20 microns.

That single Rydberg atom can then be converted to a photon, ensuring that – on average – only one photon is produced from a rubidium cloud containing hundreds of densely-packed atoms. Reliably producing a single photon with well known properties is important to several research areas, including quantum information systems.

The new technique was reported April 19 in Science Express, the rapid online publication of the journal Science. The research was supported by the National Science Foundation (NSF), and by the Air Force Office of Scientific Research (AFOSR).

“We are able to convert Rydberg excitations to single photons with very substantial efficiency, which allows us to prepare the state we want every time,” explained Alex Kuzmich, a professor in the School of Physics at the Georgia Institute of Technology. “This new system offers a fertile area for investigating entangled states of atoms, spin waves and photons. We hope this will be a first step toward doing a lot more with this system.”

Kuzmich and co-author Yaroslav Dudin, a graduate research assistant, have been studying quantum information systems that rely on mapping information from atoms onto entangled pairs of photons. But the Raman scattering technique they have been using to create the photons was inefficient and unable to provide the number of entangled photons needed for complex systems.

Georgia Tech graduate student Yaroslav Dudin and professor Alex Kuzmich (l-r) adjust optics as part of research into the production of single photons for use in optical quantum information processing and the study of certain physical systems. (Click image for high-resolution version. Credit: John Toon)

“This new photon source is about a thousand times faster than existing systems,” Dudin said. “The numbers are very good for our first experimental implementation.”

To create a Rydberg atom, the researchers used lasers to illuminate a dense ensemble of several hundred rubidium 87 atoms that had been laser-cooled and confined in an optical lattice. The illumination boosted a single atom from the entire cloud into the Rydberg state. Atoms excited to the Rydberg state strongly interact with other Rydberg atoms, and under suitable conditions, modify the atomic level energies and prevent more than one atom from being transferred into this state – a phenomenon known as the Rydberg blockade.

Rydberg atoms show this strong interaction within a range of 10 to 20 microns. By limiting their starting ensemble of rubidium atoms to approximately that distance, Kuzmich and Dudin were able to ensure that no more than one such atom could form.

“The excited Rydberg atom needs space around it and doesn’t allow any other Rydberg atoms to come nearby,” Dudin explained. “Our ensemble has a limited volume, so we couldn’t fit more than one of these atoms into the space available.”

Kuzmich and Dudin have been using Rydberg atoms with a principal quantum number of approximately 100. These excited atoms are much larger – as much as a half-micron in diameter – than ground state rubidium atoms, which have a quantum number of 5 and a diameter of a few Angstroms.

Once a highly excited atom was created, the researchers used an additional laser field to convert the excitation into a quantum light field that has the same statistical properties as the excitation. Because the field was produced by a single Rydberg atom, it contained just one photon, which can be used in a variety of protocols.

For the Georgia Tech group, the next goal may be development of a quantum gate between light fields. The quantum gating of photons has been proposed and pursued by many research groups, so far unsuccessfully.

“If this can be realized, such quantum gates would allow us to deterministically create complex entangled states of atoms and light, which would add valuable capabilities to the fields of quantum networks and computing,” Kuzmich said. “Our work points in this direction.”

Beyond quantum information systems, the new single-photon system could also help scientists investigating other areas of physics.

“Our results also hold promise for studies of dynamics and disorder in many-body systems with tunable interactions,” Kuzmich explained. “In particular, translational symmetry breaking, phase transitions and non-equilibrium many-body physics could be investigated in the future using strongly-coupled Rydberg excitations of an atomic gas.”

The single-photon work complements research being done in the Kuzmich lab on long-lived quantum memories. A new Air Force Office of Scientific Research Multidisciplinary University Research Initiative (MURI) was recently awarded to a consortium of seven U.S. universities that will work together to determine the best approach for generating quantum memories based on interaction between light and matter. Georgia Tech leads the MURI.

“With this new work, we have demonstrated a new, deterministic source of single photons,” Kuzmich said. “In its first experimental realization, it already out-performs other types of single photons that have been pursued during the past decade around the world, including in our group. With further increases in efficiency and generation rate – and integration with long-lived quantum memories being developed in related work – such a single-photon source may make possible optical quantum information processing.”

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Ignacio Taboada, an assistant professor in the Georgia Tech School of Physics, poses with a schematic of the IceCube Neutrino Observatory, which is located at the South Pole. (Click image for high-resolution version. Credit: Gary Meek)

Cosmic rays are electrically charged particles, such as protons, that strike Earth from all directions, with energies up to one hundred million times higher than those created in man-made accelerators. The intense conditions needed to generate such energetic particles have focused physicists’ interest on two potential sources: the massive black holes at the centers of active galaxies, and the exploding fireballs observed by astronomers as gamma ray bursts (GRBs).

IceCube is using neutrinos, which are believed to accompany cosmic ray production, to explore these theories. In a paper published in the April 19 issue of the journal Nature, the IceCube collaboration – which includes a Georgia Institute of Technology scientist — describes a search for neutrinos emitted from 300 gamma ray bursts observed, most recently in coincidence with the SWIFT and Fermi satellites, between May 2008 and April 2010. Surprisingly, they found none – a result that contradicts 15 years of predictions and challenges one of the two leading theories for the origin of the highest energy cosmic rays.

“The result of this neutrino search is significant because for the first time we have an instrument with sufficient sensitivity to open a new window on cosmic ray production and the interior processes of GRBs,” said IceCube spokesperson and University of Maryland physics professor Greg Sullivan. “The unexpected absence of neutrinos from GRBs has forced a re-evaluation of the theory for production of cosmic rays and neutrinos in a GRB fireball and possibly the theory that high energy cosmic rays are generated in fireballs.”

IceCube is a high energy neutrino telescope at the geographical South Pole in Antarctica, operated by a collaboration of 250 physicists and engineers from the United States, Germany, Sweden, Belgium, Switzerland, Japan, Canada, New Zealand, Australia and Barbados. The IceCube Neutrino Observatory was built under a National Science Foundation (NSF) Major Research Equipment and Facilities Construction grant, with assistance from partner funding agencies around the world. The NSF Office of Polar Programs continues to support the project with a maintenance and operations grant. Construction was finished in December 2010.

This schematic shows the IceCube Neutrino Observatory located at the South Pole. The facility includes a cubic kilometer of glacial ice and 5,160 optical sensors embedded in it. (Click image for high-resolution version)

“One of the main objectives of IceCube is to search for the sources of the highest energy cosmic rays,” explained Ignacio Taboada, an assistant professor in the Georgia Tech School of Physics who has been involved in IceCube since its planning stages. “Gamma ray bursts have always been high on the list of potential sources for cosmic rays. Though not completely ruled out, the mechanisms by which GRBs could produce these cosmic rays are now significantly constrained by these results. We will keep looking for the sources, and our chances of finding them will increase as we accumulate more data to improve our sensitivity.”

IceCube observes neutrinos by detecting the faint blue light produced in neutrino interactions in ice. Neutrinos are of a ghostly nature; they can easily travel through people, walls, or the planet Earth. To compensate for the antisocial nature of neutrinos and detect their rare interactions, IceCube is built on an enormous scale. One cubic kilometer of glacial ice, enough to fit the great pyramid of Giza 400 times, is instrumented with 5,160 optical sensors embedded up to 2.5 kilometers deep in the ice.

GRBs, the universe’s most powerful explosions, are usually first observed by satellites using X-rays and/or gamma rays. GRBs are seen about once per day, and are so bright that they can be seen from half way across the visible universe. The explosions usually last only a few seconds, and during this brief time they can outshine everything else in the universe.

“Although we have not discovered where cosmic rays come from, we have taken a major step towards ruling out one of the leading predictions,” said IceCube principal investigator and University of Wisconsin – Madison physics professor Francis Halzen.

Improved theoretical understanding and more data from the compete IceCube detector will help scientists better understand the mystery of cosmic ray production. IceCube is currently collecting more data with the finalized, better calibrated, and better understood detector.

For more information about IceCube, visit www.icecube.wisc.edu.

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Image shows an assembled magnetically actuated peel test (MAPT) specimen being prepared for analysis at the Georgia Institute of Technology. The silver cylinder in the center is the permanent magnet. (Click image for high-resolution version. Image courtesy of Greg Ostrowicki and Suresh Sitaraman)

The fixtureless and noncontact technique, known as the magnetically actuated peel test (MAPT), could help ensure the long-term reliability of electronic devices, and assist designers in improving resistance to thermal and mechanical stresses.

“Devices are becoming smaller and smaller, and we are driving them to higher and higher performance,” said Suresh Sitaraman, a professor in the George W. Woodruff School of Mechanical Engineering at the Georgia Institute of Technology. “This technique would help manufacturers know that their products will meet reliability requirements, and provide designers with the information they need to choose the right materials to meet future design specifications over the lifetimes of devices.”

The research has been supported by the National Science Foundation, and was reported in the March 30, 2012 issue of the journal Thin Solid Films.

Modern microelectronic chips are fabricated from layers of different materials – insulators and conductors – applied on top of one another. Thermal stress can be created when heat generated during the operation of the devices causes the materials of adjacent layers to expand, which occurs at different rates in different materials. The stress can cause the layers to separate, a process known as delamination or de-bonding, which is a major cause of microelectronics failure.

“We need to find out if these layers will separate over time as they are used and subjected to thermal and other stresses,” Sitaraman explained. “These systems are used in a wide range of applications from cell phones and computers to automobiles, aircraft and medical equipment. They must be reliable over the course of their expected lifetimes.”

Georgia Tech School of Mechanical Engineering professor Suresh Sitaraman (left) and doctoral student Gregory Ostrowicki examine a specimen fabricated for the magnetically actuated peel test (MAPT). An enlarged image of the specimen is shown behind them. (Click image for high-resolution version. Credit: Gary Meek)

Sitaraman and doctoral student Gregory Ostrowicki have used their technique to measure the adhesion strength between layers of copper conductor and silicon dioxide insulator. They also plan to use it to study fatigue cycling failure, which occurs over time as the interface between layers is repeatedly placed under stress. The technique may also be used to study adhesion between layers in photovoltaic systems and in MEMS devices.

The Georgia Tech researchers first used standard microelectronic fabrication techniques to grow layers of thin films that they want to evaluate on a silicon wafer. At the center of each sample, they bonded a tiny permanent magnet made of nickel-plated neodymium (NdFeB), connected to three ribbons of thin-film copper grown atop silicon dioxide on a silicon wafer.

The sample was then placed into a test station that consists of an electromagnet below the sample and an optical profiler above it. Voltage supplied to the electromagnet was increased over time, creating a repulsive force between the like magnetic poles. Pulled upward by the repulsive force on the permanent magnet, the copper ribbons stretched until they finally delaminated.

Photograph shows an assembled magnetically actuated peel test (MAPT) specimen being prepared for study at the Georgia Institute of Technology. The silver cylinder in the center is the permanent magnet. (Click image for high-resolution version. Credit: Gary Meek)

With data from the optical profiler and knowledge of the magnetic field strength, the researchers can provide an accurate measure of the force required to delaminate the sample. The magnetic actuation has the advantage of providing easily controlled force consistently perpendicular to the silicon wafer.

Because many samples can be made at the same time on the same wafer, the technique can be used to generate a large volume of adhesion data in a timely fashion.

But device failure often occurs gradually over time as the layers are subjected to the stresses of repeated heating and cooling cycles. To study this fatigue failure, Sitaraman and Ostrowicki plan to cycle the electromagnet’s voltage on and off.

“A lot of times, layers do not delaminate in one shot,” Sitaraman said. “We can test the interface over hundreds or thousands of cycles to see how long it will take to delaminate and for that delamination damage to grow.”

The test station is small enough to fit into an environmental chamber, allowing the researchers to evaluate the effects of high temperature and/or high humidity on the strength of the thin film adhesion. This is particularly useful for electronics intended for harsh conditions, such as automobile engine control systems or aircraft avionics, Sitaraman said.

“We can see how the adhesion strength changes or the interfacial fracture toughness varies with temperature and humidity for a wide range of materials,” he explained.

Georgia Tech School of Mechanical Engineering professor Suresh Sitaraman (left) and doctoral student Gregory Ostrowicki examine a specimen scheduled for analysis using the magnetically actuated peel test (MAPT). An enlarged image of the specimen is shown behind them. (Click image for high-resolution version. Credit: Gary Meek)

So far, Sitaraman and Ostrowicki have studied thin film layers about one micron in thickness, but say their technique will work on layers that are of sub-micron thickness. Because their test layers are made using standard microelectronic fabrication techniques in Georgia Tech’s clean rooms, Sitaraman believes they accurately represent the conditions of real devices.

“To get meaningful results, you need to have representative processes and representative materials and representative interfaces so that what is measured is what a real application would face,” he said. “We mimic the processing conditions and techniques that are used in actual microelectronics fabrication.”

As device sizes continue to decline, Sitaraman says the interfacial issues will grow more important.

“As we continue to scale down the transistor sizes in microelectronics, the layers will get thinner and thinner,” he said. “Getting to the nitty-gritty detail of adhesion strength for these layers is where the challenge is. This technique opens up new avenues.”

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Writer: John Toon