Researchers around Jianhua Hao demonstrate targeted in vivo imaging with a new emission probe. The fluorescent molecule has a bright and relatively small band emission around 1330nm and is rapidly excreted by a life organism making it very suitable for clinical NIR-II imaging applications.
The researchers show imaging of kidney function and filling of a mouse bladder as well as detection of small millimeter sized tumors. Furthermore, the researchers can combine their technique with X-ray imaging as the probe is a good X-ray imaging agent as well.
Confocal microscopes are widely used in physical and in particular life sciences as they offer higher resolution as standard widefield imaging. If high acquisition speeds and frame rates need to be achieved typically scanning confocal systems are used that permit to scan a field of view with an excitation light source with kHz rates
The research group of Ardemis Boghossian from Lausanne, Switzerland has just published a report about their research in collaboration with Nikon and Crest Optics to transfer scanning disc confocal microscopy to the SWIR/NIR-II wavelength range using a Princeton Instruments NIRvana-ST camera for detection.
SWIR imaging is increasingly established in life science due to low absorption and scattering of tissue in this wavelength band. The researchers show with several application examples that the new setup increases the spatial resolution while still allowing for image acquisition at high frame rates. For example, carbon nanotubes can be precisely localized in chloroplasts, monitoring nanoparticle movement in solvents allows to measure diffusivities and using the high vertical resolution allows for glucose concentration measurements using nanosensors at different depth of a sample. This research lays the groundwork for establishing and expanding this technique to monitor the location and movement of nanoparticles and sensors.
Hiromichi Kataura from the National Institute of Advanced Science and Technology in Japan is well known in the scientific community for his research on physics and applications of carbon nanotubes. Recently him and his team are using carbon nanotubes as small bio probes to image physiological and biochemical activity in-vivo.
Specifically, the researchers are looking at brown adipose tissue (BAT). BAT plays an important role in the regulation of body temperature and when stimulated, anti-obesity and diabetic effects are increased. A deep understanding of the mechanisms in BAT is important for the development of drugs to fight metabolic syndrome.
The researchers describe that the current standard method for observing BAT is positron emission tomography (PET) or histological investigation (ex-vivo). While PET is useful it does not always reflect the metabolic activity of the fat tissue well, so one of Kataura’s goals is to provide new observation methods that correctly correlate to metabolism and allow for in-vivo observations.
The team prepares carbon nanotubes emitting fluorescence light between 1000-1400 nm where light can penetrate deeply in tissue without absorption or scattering by biomolecules. This range also has low tissue autofluorescence. The nanotubes are can be highly adapted to be specific to the BAT tissue. The team shows that the fluorescence signal can be successfully applied to observe the activity in BAT tissue.
The group of Markita Landry at UC Berkely is researching methods to measure and understand the mechanisms of life on the nanoscale. Her group uses Carbon Nanotubes as a fluorescent probe on the nm length scale. Nanotubes can be built to measure different molecules in living systems, but they can also be precisely engineered to emit at certain wavelength by changing their diameter and chirality.
The group published research using nanotube species emitting light at different wavelengths that each target different molecules, multiplexing the collection of information of several biochemical processes. The nanotubes are tuned to emit light in the SWIR/NIR-II range where low absorption and scattering in tissue give the highest spatial accuracy of light emission.
Radiation therapy is one standard method for treating certain types of cancerous tumors. The challenge in all radiation therapies is always to precisely localize the diseased tissue and deliver a lethal radiation dose at exactly that location without damaging surrounding healthy tissues. Brian Pogues group from Dartmouth in Vermont has pioneered the use of gated ICCD cameras to detect Cherenkov light that gets produced by ionizing radiation in matter. Measuring the Cherenkov emission allows this group to measure radiation deposited at certain locations within a human body.
This technology is currently commercialized by a company spun off from Prof. Pogues group. While Cherenkov ligh is mostly emitted in the blue to visible region, SWIR light gives much higher accuracy about the position of radiation in tissue due to the low scattering and absorption by tissue. The researchers are now reporting on using fluorescent quantum dots that are distributed in a tissue sample and designed to absorb the Cherenkov radiation and emit SWIR radiation. The group is now working on applying this technique in radiation therapy as well as imaging molecular processes with targeted nanoscale probes.
High-resolution spectroscopy for measuring fine structures and spin interactions in quantum materialsCustomer Stories
Dr. Jörg Debus
TU Dortmund University, Germany
Jörg Debus and his team at TU Dortmund University in Germany conduct research on materials with high potential for applications in optical quantum information processing, spintronics as well as quantum sensing. The group studies structures, such as quantum dots, as well as 2D materials, semiconducting defects in diamond and rare-earth ions quantum wells. For optically driven spintronics and quantum information processing, the coherent spin manipulation with ultrashort laser pulses requires knowledge about the fine structures of excitons, in particular, the electron and hole g-factors: they define the frequency of quantum bits. In addition to the spin level structures, the interactions between confined carriers play a crucial role, since they limit the quantum information handling due to spin relaxation.
One example of experiments performed by the Debus team are recent measurements on the energy and spin structure of nitrogen vacancies in diamond crystals. Due to their unique electron confinement, the electron spins demonstrate robust coherence times exceeding several seconds at room temperature that are suitable for quantum information and quantum sensing applications. For such use it is important to know the fine structure of energy levels corresponding to different spin states in magnetic fields and understand the interaction mechanisms of charge carriers in the material. The Debus team uses optical spectroscopy to measure these properties and to spectrally resolve the fine structures.
In addition to photoluminescence spectroscopy, another technique used by the Debus team is spin-flip Raman scattering, a process similar to ordinary Raman scattering, however instead of a different phonon state the initial and final material states have different spin properties. The spin-flip signal is detected at a position that is spectrally shifted from the excitation laser line by the energy difference of the spin states. Spin-flip Raman scattering as a resonant process is not only useful for measuring spin levels, but also to prepare carriers confined in quantum dots in specific spin states. Above all, the scattering mechanism helps to identify the spin interaction between electrons and holes. Most experiments in the lab are performed at cryogenic temperatures in magnetic fields, while precisely controlling the energy and polarization of the excitation light.
Figure 1: Top: Scheme of the TriVista setup used for spin-flip Raman scattering. Bottom, left: Raman spectrum shows the spin-flip signal of electrons confined in InGaAs/GaAs quantum dots taken at a magnetic field of 8 T and a temperature of 6 K. The excitation was done at 1.39 eV (892 nm), the detection was provided by a LN2-cooled Spec-10 CCD camera. Bottom right: Scheme of the electron spin flip (blue colored) for resonant σ- circular-polarized excitation of an exciton in oblique magnetic field geometry.
Research in the Debus lab does not only focus on one, but on a broad range of materials. The spectroscopy system needs to adapt to changing signal wavelength and use of lasers at different excitation wavelength or tunable lasers as well as obtain sufficiently high spectral power to resolve the details of the fine structure and interactions of spin states which are tuned by an applied magnetic field.
One problem in resolving the fine structure of, for example, semiconductor quantum dots is inhomogeneous broadening due to a distribution of energy levels caused by small variations in size and shape of the quantum dots. By tuning the excitation wavelength in resonance with specific quantum dot states, the signal of other dots in the sample is suppressed reducing spectral broadening. However, laser excitation will be in close spectral vicinity of the detection signal. The same is true for signals in resonant spin-flip Raman scattering which are slightly shifted by a fraction of an meV (a few cm-1) from the excitation laser line.
Spectroscopic measurements in close vicinity to the laser line are extremely challenging. The intensity of elastic scattered light is often much stronger than the signal and interfere with the detection of weak signals on the detector. Filters have to be used to reduce the amount of laser light before detection. The filters need sharp cutoffs and transitions width to measure close to the laser line, however changing excitation wavelength typically requires use or purchase of additional filters.
The TriVista allows us to perform challenging optical spectroscopy with high resolution as close as a few 100 μeV (0.8 cm-1) from the excitation laser line
The Debus team uses aTriVista TR555 triple stage spectroscopy system to achieve not only high resolution and strong stray-light suppression (for signals close to the laser line), but also flexibility to adapt to the changing experimental requirements such as different materials, excitation and detection wavelengths. Moreover, the intensity of the spin-flip Raman scattering signals is quite low so that a high efficiency of the optical components and detector sensitivity are necessary.
TriVista systemsare built from 3 spectrometer stages. By combining the diffractive power of all 3 stages high spectral resolution is achieved up to 300% higher compared to a single stage. Another operating mode combining all 3 stages allows for recording signals in the vicinity of the laser line as close as 5 cm-1 (0.62 meV) using a CCD camera for detection. In this mode of operation, the first 2 stages are linked together in a way to act as a signal bandpass filter that is spectrally dispersed by the third stage. Alternatively, the Debus team sometimes uses the high- resolution mode with a single channel detector (e.g. PMT) for detection. It also achieves high stray-light rejection and is suitable for measurements that do not require the spectral multiplexing advantage of detection using a CCD.
TheTriVista system easily adapts to the changing experimental requirements of the lab. TriVista can be quickly adapted to any laser or signal wavelength from the UV to the IR without use of additional edge or notch filters that need to be changed for each operating wavelength.
TriVista systems also operate up to 4 signal output ports (one on stage 1 and stage 2, two on stage 3) and additionally to the combined operation modes described above, each stage can be operated independently of each other. The Debus team uses efficient detectors for visible and IR detection on different output ports as well as intensified CCD cameras such as thePI-MAXfor time-resolved measurements with nanosecond resolution.
Having the capability of high resolution, high stray-light reduction measurements at any wavelength as well as multiple efficient detection and operation options allows the TriVista systems to fulfil the multiple needs and requirements for all aspects of new and quantum material research by the Debus team.
Researchers around Brian Pogue and Scott Davis from Dartmouth College in the US have published a conference report about their experiments on using 2D imaging for singlet oxygen imaging and dosimetry in cancer treatment by radiation therapy. Specifically, they perform photodynamic therapy where the photosensitizer molecule used triggers a destructive chain reaction in tumor cells on radiation. As singlet oxygen (an excited form of the oxygen molecule) is closely involved in this reaction, direct detection of emission from this molecule can be helpful in monitoring the reaction dose as well as estimating the deposited energy dose during treatments.
Highly sensitive, deeply cooled 2D InGaAs cameras are very well suited to monitor the emission of singlet oxygen. In the current study the researchers have developed several setups and they included a Princeton Instruments NIRvana to further increase the detection sensitivity. They performed extensive tests to show that the setup is very suitable for sensitive dosimetry and will continue with extensive in vivo and animal studies. This research should be extremely relevant for all researchers working in bioimaging as well as cancer research and treatment.
Bioimaging in the NIR-II/SWIR wavelength range is currently used to research cancer detection and treatment, monitoring metabolic and organ functions or measuring blood flow and heartrate non-invasively. The research is currently progressing from early proof of concept stages to describe more specific bio experiments using the imaging as a tool to monitor the systems. However, research to find more suitable and functional nanoprobes, small molecules and particles that emit fluorescent light in the SWIR, with the potential to be functionalized and used in vivo, is still a very important part of this research.
A research team from China is reporting on experiments using particles based on rare earth metals and they can show high performance and tunability of these probes. The functionality is shown by sensitive in vivo measurements of tumors and blood vessels. The researchers note that these probes are promising due to their bright emission as well as biocompatibility.
Development and testing of new molecular/nano-bio probes is an important part of NIR-II/SWIR biological imaging and diagnostics. Research in particular is focused on finding probes of high efficiency that can be used in human/clinical applications. Previously the lab of Moungi Bawendi at MIT successfully demonstrated use of a fluorophore called indocyanine green (ICG) which is clinically approved but does not optimally emit radiation in the desired wavelength range
In a recent article in Nature Communications researchers around Fan Zhang from Fudan University (Shanghai, China) developed molecules similar to ICG but optimized to emit in the NIR-II/SWIR wavelength range while still exhibiting low toxicity for potential clinical applications. The researchers demonstrate imaging up to 8mm into tissue as well as reliable, non-invasive measurements of pH values in mouse stomachs. Having demonstrated the basic use of this class of fluorophores opens up the path to more wider applications in biomedical sensing.
Several analytical methods have been commonly employed for carbon nanomaterial detection including electron microscopy (TEM/SEM), Raman spectroscopy, and mass spectrometry. However, each has limitations, for example a lack of spatial resolution specificity for differentiating carbon nanomaterials from the carbon in the biomass of living organisms. Near IR fluorescence imaging has emerged as promising tool for detection and quantification of singlewalled carbon nanotubes (SWCNTs) for in vivo studies.
Collaborating researchers from the University of Florida, University of Texas and Duke University in the US published a study demonstrating that near infrared fluorescence was an effective method for pulmonary retention of SWCNTs in a murine model. They used a custom imaging system based on detection with a Princeton Instruments OMA V, 2D InGaAs array for capturing the emission from carbon nanotubes. The concentration of nanotubes is deliberately chosen to be at or below the detection threshold of other technologies such as Raman spectroscopy. The experiments use pristine, unaltered and not functionalized nanotubes. The methods are also applied on tracking and quantifying concentration of carbon nanotubes in lung tissues to assess pulmonary retention and distribution leading to potential health effects in mice.