QCL-based glucose testing, hyperspectral imaging and molecular techniques lead the list of the year’s most popular stories. The brain is having a big year. At the end of September, the National Photonics Initiative launched the Photonics Industry Neuroscience Group, an industry consortium that includes several well-known photonics companies. The industry leaders plan to invest upwards of $30 million for photonics research and development in support of the White House’s BRAIN Initiative. Targeted technologies include the following: imaging optics, laser sources, automated scanning technology and high-resolution cameras; miniature, affordable, portable and implantable microscopes for therapeutic screening; fluorescent-protein engineering technology; and automated software for detailed mapping of the human brain. Superresolution STED microscopy (circular inset image) provides images approximately 10 times sharper than a conventional light microscope (outer image) can achieve. Courtesy of the Max Planck Institute for Biophysical Chemistry. Biophotonics itself is having a big year, too – and the biggest news of all may have come with the announcement of the Nobel Prize in chemistry, which honored the work of superresolution microscopy pioneers Dr. Eric Betzig, Dr. Stefan W. Hell and Dr. W.E. Moerner. The techniques these three developed have enabled scientists to see how molecules create synapses between nerve cells in the brain; track proteins involved in Parkinson’s, Alzheimer’s and Huntington’s diseases; and follow individual proteins in fertilized eggs as they divide into embryos. “Today, nanoscopy is used worldwide, and new knowledge of greatest benefit to mankind is produced on a daily basis,” wrote the Royal Swedish Academy of Sciences, which presents the Nobels. As exciting as those two announcements are, they are by no means the only biophotonics news released in 2014. Here’s a look back at some of the other big accomplishments that the biophotonics research community has announced over the past year. These stories represent Photonics.com’s most-shared laser, imaging, microscopy and spectroscopy headlines of 2014. QCL Tests Blood Sugar Diabetic patients currently endure finger pricks multiple times a day to help monitor their blood sugar levels, but a blood-sugar test incorporating an IR quantum cascade laser could spare them the pain, Princeton University researchers announced in July (doi: 10.1364/boe.5.002397). When directed at the patient’s palm, the laser light is partially absorbed by sugar molecules in the body; the amount of absorption is used to measure the level of blood sugar. Instead of blood, the light targets dermal interstitial fluid, which strongly correlates with blood sugar levels. Mid-IR wavelengths need relatively high power and stability to penetrate the skin; the quantum cascade laser produces the desired frequency, power and stability. Courtesy of Frank Wojciechowski/Princeton University. The new laser system originally filled up a moderate-sized workbench, the researchers said, and also needed an elaborate cooling system to function properly. But the laser now operates at room temperature. Next, they plan to shrink the laser system to a portable size, which would help diabetics to check their own condition. “We are working hard to turn engineering solutions into useful tools for people to use in their daily lives,” said Dr. Claire Gmachl, a professor of electrical engineering at Princeton. Hyperspectral Imaging Reduces Amputation Threat Hyperspectral imaging (HSI) can help surgeons make quick decisions in the operating room, as it detects issues not visible to the naked eye. Dallas surgeon Dr. Javier La Fontaine, associate professor of plastic surgery at the University of Texas Southwestern Medical Center, explained at Photonics West 2014 that using this imaging technique can reduce the threat of amputation. “It has saved some patients time spent in returns to the operating room,” he said. “It lets us more comfortably address the severity of the wound infection.” La Fontaine cited a case in which HSI helped save a patient’s injured toe, which he initially thought would require amputation. HSI was used to monitor oxygen levels in the tissues, which provided instant analysis of this patient’s foot. “If you take too little, the wound can get reinfected,” La Fontaine said. “This can lead to a return to the operating room and can lead to a greater level of amputation.” The HSI instrument used a Texas Instruments digital micromirror device, a set of millions of tiny mirrors each measuring about 1 µm. The device generates high-resolution images to indicate the presence of certain molecules. Molecular Technique Enables Live-Cell Imaging A new optical imaging platform represents a significant step toward a long-standing scientific goal: visualizing small biomolecules inside living biological systems with minimum disturbance. A team at Columbia University in New York announced in March a method (doi: 10.1038/nmeth.2878) to image a broad spectrum of small biomolecules, such as molecular drugs and nucleic acids, amino acids and lipids, viewing where they are localized and how they function inside cells. Courtesy of Columbia University. “Our new technique will open up numerous otherwise-difficult studies on small biomolecules in live cells and animals,” said Wei Min, an assistant professor of chemistry at Columbia. “In addition to basic research, our technique could also contribute greatly to translational applications.” The researchers coupled the laser-based technique stimulated Raman scattering (SRS) microscopy with a small but highly vibrant alkyne tag (a chemical bond that, when stretched, produces a strong Raman scattering signal at a frequency different from natural molecules inside cells). The technique labels the small molecules with the tiny alkyne tag and obtains high detection specificity and sensitivity by SRS imaging. The unique stretching motion of the carbon-carbon bond that is carried by the small molecules produces a 3-D map of them inside living cells and animals. Optical Brain Monitoring for Better Stroke Care Advanced technology and simple body positioning could be key in providing acute stroke patients with more effective individualized treatment in real time. A team from the University of Pennsylvania in Philadelphia announced in March a new optical device that can noninvasively and continuously monitor cerebral blood flow (CBF) in stroke patients to gauge how their body positioning could impact blood flow to the brain. When stroke patients are first admitted to the hospital, they typically are kept flat for 24 hours to increase CBF in the brain regions surrounding the damaged tissue. This position helps most stroke patients, but the new device has shown that, for 29 percent of patients, an elevated rather than flat head position increases CBF and is therefore more beneficial. The new technology, called diffuse correlation spectroscopy, uses a noninvasive probe to measure fluctuations of NIR light that has traveled through the skull and into the brain, then back out to the tissue surface. These fluctuations, caused by moving red blood cells in tissue, have been shown to accurately track CBF in underlying brain tissue. The device could someday be used routinely for all patients with acute brain injury. Light-ActivatedNneurons Could Restore Paralyzed Muscles A light-based method could help paralyzed muscles move again. Unveiled in April by University College London and King’s College London, the technique potentially could restore the function of muscles afflicted by motor neuron disease or spinal cord injury. “This strategy has significant advantages over existing techniques that use electricity to stimulate nerves, which can be painful and often results in rapid muscle fatigue,” said Dr. Linda Greensmith of UCL. “If the existing motor neurons are lost due to injury or disease, electrical stimulation of nerves is rendered useless as these, too, are lost.” Courtesy of Dr. Barney Bryson, University College London Institute of Neurology. The new method, which has been tested on mice (doi: 10.1126/science.1248523), transplants motor neurons created from stem cells into injured nerve branches. “We custom-tailored embryonic stem cells so that motor neurons derived from them can function as part of the muscle pacemaker device,” said Dr. Ivo Lieberam of King’s College. These cells are equipped with a molecular light sensor, allowing the researchers to control the motor neurons with flashes of blue light. They could adjust muscle control by modifying the intensity, duration and frequency of the light pulses. The researchers expect to conduct human trials within the next five years. Biometric Watches Noninvasively Monitor Vital Signs The ability to monitor vital signs is moving out of the clinic. In May, researchers in Israel and the Netherlands published articles on two noninvasive, wearable sensing devices that can be worn like a wristwatch to monitor biometrics. The sensors detect changing patterns of scattered light – one tracks a person’s glucose concentration and hydration levels (doi: 10.1364/BOE.5.001926), and the other monitors pulse (doi: 10.1364/BOE.5.002145). “Around 96 percent of our in vivo measurements were within a range of 15 percent deviation from the readout of a medical reference glucometer device,” said Dr. Zeev Zalevsky of Israel’s Bar-Ilan University. “The main factor for errors now is the stability of our device on the wrist of the user.” The sensor detects changes in glucose concentration by analyzing varying patterns in back-scattered light produced by laser-generated wavefronts of light that illuminate skin on the wrist near an artery; a camera measures the changes over time. “Glucose is the holy grail of the world of biomedical diagnostics, and dehydration is a very useful parameter in the field of wellness, which is one of our main commercial aims,” Zalevsky said. Handheld Scanner Roots Out Brain Tumor Traces A new Raman laser scanner could help surgeons more effectively remove cancerous brain tumors. A team at the Memorial Sloan Kettering Cancer Center in New York is developing the handheld device, which could be used to root out malignant cells during surgery so that fewer or none are left behind to form new tumors. Conventional methods are not accurate enough to identify all of the cancerous cells that need to be excised. In a study published in August (doi: 10.1021/nn503948b), the researchers evaluated the ability of the Raman scanner – guided by surface-enhanced Raman scattering nanoparticles injected earlier into the mice – to identify the microscopic tumor extent in a genetically engineered RCAS/tv-a glioblastoma mouse model. The handheld scanner was tested alongside a stationary Raman imaging device. The two complemented each other, the researchers said, although the new scanner allowed nearly real-time scanning and detected additional microscopic foci of cancer not detected by the stationary imager. This device has the potential to move readily into clinical trials, the researchers said, and surgeons might be able to use it in the future to treat other types of brain cancer. Microscopy Enables Detailed Insights Into Mitochondria A microscopy technique published in April combines confocal and two-photon excitation microscopy with in situ pharmacological and genetic manipulation to give mitochondrial-level insight into the nervous system. The new method allows researchers to record the oxidation states of individual mitochondria with high spatial and temporal resolution, and to analyze how reduction/oxidation (redox) signaling unfolds in single cells and organelles in real time. The nerve-cell mitochondria were imaged with a fluorescent redox sensor. Shown here is a peripheral nerve with the neuromuscular end plates stained in red. Courtesy of Ludwig Maximilian University and the Technical University of Munich. “Redox signals have important physiological functions, but can also cause damage, for example, when present in high concentrations around immune cells,” said professor Dr. Martin Kerschensteiner of Ludwig Maximilian University of Munich and SyNergy, the Munich cluster for Systems Neurology. His team collaborated with that of Technical University of Munich professor Dr. Thomas Misgeld, also of SyNergy. The researchers used redox-sensitive variants of green fluorescent protein as visualization tools. They combined these with other biosensors and dyes, which allowed redox signals and mitochondrial calcium currents to be monitored simultaneously, along with changes in electrical potential and pH gradient across the mitochondrial membrane. The method allowed them to study, for the first time, redox signal induction in response to neural damage in the mammalian nervous system – in this case, spinal cord injury (doi: 10.1038/nm.3520). Enhanced CARS Enables High-Speed Tissue Imaging An enhanced spectroscopy technique for analyzing biological cells and tissues delivers stronger signals than those attained through conventional practices. A team from the National Institute of Standards and Technology in Gaithersburg, Md., in conjunction with the Cleveland Clinic, announced in July that it had demonstrated the improved method based on characteristic molecular vibration signatures (doi: 10.1038/nphoton.2014.145). A false-color BCARS image of mouse liver tissue (left) picks out cell nuclei in blue, collagen in orange and proteins in green. The image of mouse tumor and normal brain tissue (right) has been colored to show cell nuclei in blue, lipids in red and red blood cells in green. The images show an area about 200 µm across. Courtesy of Dr. Charles Camp Jr./NIST. The technique, called broadband coherent anti-Stokes Raman scattering (BCARS), can deliver signals 10,000 times stronger than those obtained through spontaneous Raman scattering and is also 100 times stronger than comparable coherent Raman scattering techniques, using a much larger portion of the vibrational spectrum. The technique accesses the same spectral region in which existing coherent Raman methods work, but it also picks up signals from the “fingerprint” spectral region, which contains most of the useful molecular identification information. The new instrument uses excitation light quickly and efficiently; conventional coherent Raman instruments must tune two separate laser frequencies to excite and read different Raman vibration modes in the sample. It is fast and accurate enough for the creation of high-resolution images of biological specimens that contain detailed spatial information on specific biomolecules. Noninvasive Brain Control Possible With New Opsins Thanks to a new molecule, optogenetics has put brain control in the hands of scientists. A new protein, Jaws, was developed at MIT in Cambridge, Mass.; the study was published in July (doi: 10.1038/nn.3752). It is sensitive to red light and enables neurons to be manipulated noninvasively; it also allows a larger volume of tissue to be influenced simultaneously. In experiments, the researchers essentially shut down neural activity in the brains of mice using a light source outside the skull. This suppression of activity penetrated as deeply as 3 mm into the brain. Courtesy of Jose-Luis Olivares/MIT. The new technique was found to be as effective as existing neural silencers, which use other wavelengths and rely on invasive methods such as implanted optical fibers. The researchers had identified two light-sensitive chloride ion opsins as a possibility because they respond to red light, which has been shown to penetrate deeper into living tissue than blue or green light. However, the researchers found that these molecules did not produce enough photocurrent to control neuron activity. To overcome this hurdle, Amy Chuong, a graduate student at MIT, engineered a relative of chloride ion. The new protein retains sensitivity to red light but also features a stronger photocurrent. “This exemplifies how the genomic diversity of the natural world can yield powerful reagents that can be of use in biology and neuroscience,” said optogenetics pioneer Dr. Edward Boyden of MIT. Other Popular BioPhotonics Stories of 2014 Adaptive Optics Enhances Subcellular Imaging: www.photonics.com/a56074 Nanoballoons and Lasers: A New Cancer Fighter: www.photonics.com/a56025 Switching an Antibiotic On and Off with Light: www.photonics.com/a55994 Optogenetic Method Erases, Reactivates Memories: www.photonics.com/a56267 Light Mitigates Alzheimer’s Symptoms: www.photonics.com/a56280 3-D Cell Imaging Through Microfluidic Channels: www.photonics.com/a56000 Carbon Spaser Could Zap Cancer Cells: www.photonics.com/a56106 Nano-Optical Tweezers Make Single-Molecule Protein Study Easier: www.photonics.com/a56244 Laser Triggers Stem Cells to Fix Teeth: www.photonics.com/a56256 Laser Process Shows Promise for Skin Cancer Treatment: www.photonics.com/a55818 Optical Scanner Shines New Light on Brain: www.photonics.com/a56229 Brain Mapping Seeks New Paths: www.photonics.com/a55937 Optogenetics Allows Light to Switch Off Anxiety: www.photonics.com/a56053 Fluorescence Sensor Diagnoses Diabetes: www.photonics.com/a56483 SERS Advances Noninvasive Prostate Cancer Screening: www.photonics.com/a56622