3D Spatial Resolution Research Articles

The Influence of Size, Shape, and Twin Boundaries on Heat‐Induced Alloying in Individual Au@Ag Core–Shell Nanoparticles

Environmental conditions during real-world application of bimetallic core-shell nanoparticles (NPs) often include the use of elevated temperatures, which are known to cause elemental redistribution, in turn significantly altering the properties of these nanomaterials. Therefore, a thorough understanding of such processes is of great importance. The recently developed combination of fast electron tomography with in situ heating holders is a powerful approach to investigate heat-induced processes at the single NP level, with high spatial resolution in 3D. In combination with 3D finite-difference diffusion simulations, this method can be used to disclose the influence of various NP parameters on the diffusion dynamics in Au@Ag core-shell systems. A detailed study of the influence of heating on atomic diffusion and alloying for Au@Ag NPs with varying core morphology and crystallographic details is carried out. Whereas the core shape and aspect ratio of the NPs play a minor role, twin boundaries are found to have a strong influence on the elemental diffusion.

Intercomparison of UAV platforms for mapping snow depth distribution in complex alpine terrain

Unmanned Aerial Vehicles (UAVs) offer great flexibility in acquiring images in inaccessible study areas, which are then processed with stereo-matching techniques through Structure-from-Motion (SfM) algorithms. This procedure allows generating high spatial resolution 3D point clouds. The high accuracy of these 3D models allows the production of detailed snow depth distribution maps through the comparison of point clouds from different dates. In this way, UAVs allow monitoring of remote areas that were not achievable previously. The large number of works evaluating this novel technique has not, to date, conducted a systematic evaluation of concurrent snowpack observations with different UAV devices. Taking into account this, and also bearing in mind that potential users of this technique may be interested in exploiting ready-to-use commercial devices, we conducted an evaluation of the snow depth distribution maps with different commercial UAVs. During the 2018–19 snow season, two multi-rotors (Parrot Anafi and DJI Mavic Pro2) and one fixed-wing device (SenseFly eBee plus) were used on three different dates over a small test area (5 ha) within Izas Experimental Catchment in the Central Pyrenees. Simultaneously, snowpack distribution was retrieved with a Terrestrial Laser Scanner (TLS, RIEGL LPM-321) and was considered as ground truth. Three different georeferencing methods (Ground Control Points, ICP algorithm over snow-free areas and RTK-GPS positioning) were tested, showing equivalent performances under optimum illumination conditions. Additionally, for the three acquisition dates, both multi-rotors were flown at two distinct altitudes (50 and 75 m) to evaluate impact on the obtained snow depth maps. The evaluation with the TLS showed an equivalent performance of the two multi-rotors, with mean RMSE below 0.23 m and maximum volume deviations of less than 5%. Flying altitudes did not show significant differences in the obtained maps. These results were obtained under contrasted snow surface characteristics. This study reveals that under good illumination conditions and in relatively small areas, affordable commercial UAVs provide reliable estimations of snow distribution compared to more sophisticated and expensive close-range remote sensing techniques. Results obtained under overcast skies were poor, demonstrating that UAV observations require clear-sky conditions and acquisitions around noon to guarantee a homogenous illumination of the study area.

Three-Dimensional Magnetic Resonance Fingerprinting in Neonates: Quantifying Regional Difference and Maturation in the Brain.

Magnetic resonance fingerprinting (MRF) allows the simultaneous measurement of multiple tissue properties in a single acquisition. Three-dimensional (3D) MRF with high spatial resolution can be used for neonatal brain imaging. The aim of this study is to apply 3D MRF to neonates and show regional differences and maturation in the brain. In this prospective study, 3D MRF using hybrid radial-interleaved acquisition was performed on phantoms and neonates from December 2019 to October 2020. For the reconstruction of 3D MRF, singular value decomposition was applied to reduce reconstruction time, and the iterative reconstruction technique was applied to improve image quality. The accuracies of T1 and T2 values derived from 3D MRF were evaluated in a phantom experiment. Regional T1 and T2 values were obtained from neonates' brain T1 and T2 maps derived from 3D MRF. Regional T1 and T2 values were compared, and their changes according to corrected gestational age were evaluated. The acquisition time for 3D MRF with a spatial resolution of 0.7 × 0.7 × 2 mm3 was less than 5 minutes. The phantom study showed high correlation between T1 and T2 values derived from 3D MRF and those from conventional spin echo sequences (T1, R2 = 0.998, P < 0.001; T2, R2 = 0.998, P < 0.001). Three-dimensional MRF was performed in 25 neonates (15 boys, 10 girls; median corrected gestational age, 263 days; interquartile range, 10 days). In neonates, T1 and T2 values differed in the frontal (median [interquartile range], 2785 [2684-2888] milliseconds and 189.8 [176.7-222.9] milliseconds), parietal (2849 [2741-2950] milliseconds and 191.6 [167.5-232.9] milliseconds), and occipital white matter (2621 [2513-2722] milliseconds and 162.9 [143.5-186.1] milliseconds), showing lower values in occipital white matter (P < 0.001). Regional T1 values showed a negative relationship with corrected gestational age (coefficient, -0.775 to -0.480; P < 0.05). Fast and high spatial resolution 3D MRF was applied to neonates. T1 and T2 maps derived from 3D MRF enabled the quantification of regional differences and maturation in the neonatal brain.

Bulk and Short‐Circuit Anion Diffusion in Epitaxial Fe2O3 Films Quantified Using Buried Isotopic Tracer Layers

AbstractSelf‐diffusion is a fundamental physical process that, in solid materials, is intimately correlated with both microstructure and functional properties. Local transport behavior is critical to the performance of functional ionic materials for energy generation and storage, and drives fundamental oxidation, corrosion, and segregation phenomena in materials science, geosciences, and nuclear science. Here, an adaptable approach is presented to precisely characterize self‐diffusion in solids by isotopically enriching component elements at specific locations within an epitaxial film stack, and measuring their redistribution at high spatial resolution in 3D with atom probe tomography. Nanoscale anion diffusivity is quantified in a‐Fe2O3 thin films deposited by molecular beam epitaxy with a thin (10 nm) buried tracer layer highly enriched in 18O. The isotopic sensitivity of the atom probe allows precise measurement of the initial sharp layer interfaces and subsequent redistribution of 18O after annealing. Short‐circuit anion diffusion through 1D and 2D structural defects in Fe2O3 is also directly visualized in 3D. This versatile approach to study precisely tailored thin film samples at high spatial and mass fidelity will facilitate a deeper understanding of atomic‐scale diffusion phenomena.

Assessment of DSM Based on Radiometric Transformation of UAV Data.

Unmanned Aerial Vehicle (UAV) is one of the latest technologies for high spatial resolution 3D modeling of the Earth. The objectives of this study are to assess low-cost UAV data using image radiometric transformation techniques and investigate its effects on global and local accuracy of the Digital Surface Model (DSM). This research uses UAV Light Detection and Ranging (LIDAR) data from 80 m and UAV Drone data from 300 and 500 m flying height. RAW UAV images acquired from 500 m flying height are radiometrically transformed in Matrix Laboratory (MATLAB). UAV images from 300 m flying height are processed for the generation of 3D point cloud and DSM in Pix4D Mapper. UAV LIDAR data are used for the acquisition of Ground Control Points (GCP) and accuracy assessment of UAV Image data products. Accuracy of enhanced DSM with DSM generated from 300 m flight height were analyzed for point cloud number, density and distribution. Root Mean Square Error (RMSE) value of Z is enhanced from ±2.15 m to ±0.11 m. For local accuracy assessment of DSM, four different types of land covers are statistically compared with UAV LIDAR resulting in compatibility of enhancement technique with UAV LIDAR accuracy.

Spatial dependency and the role of local susceptibility for velocity selective arterial spin labeling (VS‐ASL) relative tagging efficiency using accelerated 3D radial sampling with a BIR‐8 preparation

Velocity selective arterial spin labeling (VS-ASL) is a promising approach for non-contrast perfusion imaging that provides robustness to vascular geometry and transit times; however, VS-ASL assumes spatially uniform tagging efficiency. This work presents a mapping approach to investigate VS-ASL relative tagging efficiency including the impact of local susceptibility effects on a BIR-8 preparation. Numerical simulations of tagging efficiency were performed to evaluate sensitivity to regionally varying local susceptibility gradients and blood velocity. Tagging efficiency mapping was performed in susceptibility phantoms and healthy human subjects (N = 7) using a VS-ASL preparation module followed by a short, high spatial resolution 3D radial-based image acquisition. Tagging efficiency maps were compared to 4D-flow, B1 , and B0 maps acquired in the same imaging session for six of the seven subjects. Numerical simulations were found to predict reduced tagging efficiency with the combination of high blood velocity and local gradient fields. Phantom experiments corroborated numerical results. Relative efficiency mapping in normal volunteers showed unique efficiency patterns depending on individual subject anatomy and physiology. Uniform tagging efficiency was generally observed in vivo, but reduced efficiency was noted in regions of high blood velocity and local susceptibility gradients. We demonstrate an approach to map the relative tagging efficiency and show application of this methodology to a novel BIR-8 preparation recently proposed in the literature. We present results showing rapid flow in the presence of local susceptibility gradients can lead to complicated signal modulations in both tag and control images and reduced tagging efficiency.

Hard X-ray nano-holotomography with a Fresnel zone plate.

X-ray phase contrast nanotomography enables imaging of a wide range of samples with high spatial resolution in 3D. Near-field holography, as one of the major phase contrast techniques, is often implemented using X-ray optics such as Kirkpatrick-Baez mirrors, waveguides and compound refractive lenses. However, these optics are often tailor-made for a specific beamline and challenging to implement and align. Here, we present a near-field holography setup based on Fresnel zone plates which is fast and easy to align and provides a smooth illumination and flat field. The imaging quality of different types of Fresnel zone plates is compared in terms of the flat-field quality, the achievable resolution and exposure efficiency i.e. the photons arriving at the detector. Overall, this setup is capable of imaging different types of samples at high spatial resolution of below 100 nm in 3D with access to the quantitative phase information.

Micro-anatomy of the ear of the southern white rhinoceros (Ceratotherium simum simum).

The white rhinoceros is the largest of the five extant rhinoceros species. The population is declining rapidly because of intense poaching. However, normal anatomical descriptions in this species are lacking. The purpose of this study is to describe the osseous anatomy of the middle and inner ear of the southern white rhinoceros using micro-focus X-ray computed tomography imaging. Four temporal bones obtained from two 1-day old southern white rhinoceros preserved in 10% formalin were scanned. Tri-dimensional reconstructions were obtained and volumes of the middle ear ossicles and inner ear structures were calculated. Excellent high spatial resolution 3D images were obtained for all samples and virtual models of the auditory ossicles and bony labyrinth were generated. Visualization of the tympanic membrane, middle ear and inner ear structures was possible in all samples. Whereas the stapes and incus had a shape similar to their human or equine counterparts, the malleus showed a unique appearance with a long rostral branch projecting latero-distally to the manubrium. The cochlea described 2 turns rostro-laterally around its axis, with a medial direction of rotation. However, identification of the soft tissue structures of the middle ear was sometimes difficult and visualization of the small structures of the membranous labyrinth was not possible using this formalin fixation and alternative techniques should be investigated. Further investigations are needed in order to provide a complete virtual model including both soft and bone tissues of this difficultly accessible region.

Quantitative µ-CT Analysis of Scale Topology Formed During Oxidation of High SiMo Cast Iron

High SiMo cast iron components in automotive exhaust systems are exposed to high-temperature oxidation over time. Quantitative analysis of formed oxide scale is therefore important for the assessment of a component durability. The brittle nature of multilayered scale and thermal stress limits capture of a true topology using traditional 2D destructive cut and polish methods. In this study, nondestructive high spatial-resolution 3D µCT analysis was performed on 2.90-mm-diameter oxidized specimens which permitted direct observation with 3.5 µm pixel resolution. The specimens were oxidized in three sequential time steps for a total 100 h at 700 °C and 800 °C in air and combustion gas atmospheres. A MATLAB-coded algorithm was used to quantify the topology, thickness variation in internal and external scale layers, and scale/metal interface unevenness. Scale topology was linked to oxidation temperature and gas atmosphere. A water vapor environment increases scale/metal interface unevenness and scale layer thickness irregularity which were related to an accelerated oxidation rate.

3D high spatial resolution visualisation and quantification of interconnectivity in polymer films.

A porous network acts as transport paths for drugs through films for controlled drug release. The interconnectivity of the network strongly influences the transport properties. It is therefore important to quantify the interconnectivity and correlate it to transport properties for control and design of new films. This work presents a novel method for 3D visualisation and analysis of interconnectivity. High spatial resolution 3D data on porous polymer films for controlled drug release has been acquired using a focused ion beam (FIB) combined with a scanning electron microscope (SEM). The data analysis method enables visualisation of pore paths starting at a chosen inlet pore, dividing them into groups by length, enabling a more detailed quantification and visualisation. The method also enables identification of central features of the porous network by quantification of channels where pore paths coincide. The method was applied to FIB-SEM data of three leached ethyl cellulose (EC)/hydroxypropyl cellulose (HPC) films with different weight percentages. The results from the analysis were consistent with the experimentally measured release properties of the films. The interconnectivity and porosity increase with increasing amount of HPC. The bottleneck effect was strong in the leached film with lowest porosity.

Value of susceptibility-weighted MR imaging (SWI) in the detection of developmental venous anomaly

BackgroundDevelopmental venous anomaly (DVA) is probably the most common anomaly of the intracranial vasculature. DVAs consist of multiple, radially oriented dilated medullary veins that converge into a transcerebral vein. Susceptibility-weighted imaging (SWI) is a high spatial resolution 3D gradient-echo MRI sequence with phase post-processing that accentuates the paramagnetic properties of blood products such as deoxyhemoglobin, intracellular methemoglobin, and hemosiderin. Its high sensitivity to hemorrhagic particles by means of susceptibility dephasing effects within the veins allow for the accurate detection, grading, and monitoring of brain venous anomalies. In this review, we evaluated the prevalence of the brain DVAs identified by SWI in many patients who had undergone magnetic resonance imaging (MRI) with contrast administration. All images were independently reviewed by two radiologists who were blinded to other MR imaging finding. It is hoped that as SWI becomes more widely available, it will provide additional diagnostic and prognostic information that will improve the care and outcome of patients with DVAs.ResultsA total of 29 DVAs were observed with its prevalence 2.8%. The DVA caputs had mostly deep localization in about 44.8% of our DVA cases. SWI proved excellent demonstration of DVAs with the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were significantly higher than the other non-contrast MR sequences. On SWI, DVA is delineated as a signal void lesion with the normal cerebral veins.ConclusionSusceptibility-weighted imaging (SWI) is valuable in the diagnosis of DVA and should be included in routine MR assessment of the brain.

Optical coherence tomography as a noninvasive 3D real time imaging tool for the rapid evaluation of phenotypic variations in insect embryonic development.

Noninvasive visualization of embryos at different development stages is crucial for the understanding of the basic developmental biology. It is therefore desirable to have an imaging tool capable of rapidly evaluating the effects of gene manipulation or genome editing in developing embryos for the studies of gene functions and genetic engineering. Here, we propose and demonstrate a novel use of optical coherence tomography (OCT) to noninvasively exam the embryonic development of the migratory locusts in real time with 3-dimensional (3D) view capability. In particular, we obtain the sufficiently high spatial resolution tomographic 2D and 3D images of live locust embryos throughout their development processes. We show that not only we are able to noninvasively observe all previously known forms of blastokinesis as an embryo develops, such as anatrepsis, katatrepsis, revolution, rotation and diapauses, and determine their precise occurring time or duration, but also discover an unreported rotation form we named "twist." In addition, with the OCT images we determined the exact occurring time of diapauses of the locusts from Tibetan plateau for the first time. Finally, we demonstrate that OCT systems can be used to rapidly capture the development defects of genetically modified embryos in which certain genes essential for embryonic development were suppressed by RNA interference. Our work shows that OCT is an enabling imaging tool with sufficient spatial resolution for the rapid evaluation of embryonic variations of small animals.

Retrieved cerebral thrombi studied by T2 and ADC mapping: preliminary results.

BackgroundRecent advances in MRI technology makes it increasingly more competitive to CT also in the field of interventions. Multi-parametric MRI offers a significant amount of data relevant for characterization of human cerebral thrombi.Patients and methodsCerebral thrombi of 17 patients diagnosed with acute stroke were acquired by mechanical thrombectomy. The thrombi were subsequently scanned using a high spatial-resolution 3D T1-weighted MRI to obtain morphological characteristics of the thrombi and also by apparent diffusion coefficient (ADC) and transversal nuclear magnetic resonance (NMR) relaxation time (T2) mapping. The MRI results were analysed for possible correlations between thrombectomy procedure parameters (recanalization time and number of passes) and MR-measurable parameters (sample-mean ADC and T2, within-sample coefficient of variation of ADC and T2, and thrombus length).ResultsBoth MRI mapping techniques enabled a good discrimination among thrombi regions of different water mobility and compaction. Within-sample coefficient of variation of ADC was found most sensitive for discrimination between the thrombi where thrombectomy procedure was performed in a single pass and those where is was performed in two or more passes (p = 0.03). Interestingly, negative correlation was found between the recanalization time and thrombus length (ρ = -0.22).ConclusionsPreliminary results of presented study shows that pretreatment MRI assessment of thrombi in stroke patients could potentially ease stroke treatment planning. In this study it is shown that within-sample coefficient of variation of ADC could serve for prediction of possible complications during thrombectomy procedures.

ESGCT 27th Annual Congress In collaboration with SETGyc Barcelona, Spain October 22–25, 2019 Abstracts

Xenotransplantation of transgenic porcine pancreatic islets offers a promising alternative source to circumvent current limitations posed by the scarcity of allogeneic donors. We have investigated the feasibility to generate tissue engineered SLA silenced islet cell clusters (ICC) to decrease xenogeneic immune responses Pancreatic islets single cell suspensions were generated by enzymatic digestion of porcine ICCs. Single cells were silenced for SLA expression by lentiviral vectors encoding for Nanoluciferase as reporter gene and for short hairpin RNAs targeting beta2-microglobulin or class II transactivator, respectively. SLA transcripts were evaluated by real-time PCR and protein levels by flow cytometry and fluorescence microscopy analyses. The effect of SLA silencing was evaluated in human T and NK cell cytotoxicity assays. SLA-silenced pancreatic beta-cells were then reassembled into ICCs in stirred bioreactors. SLA class I silencing reached a level of up to 84% and class II by up to 50% on pancreatic islet cell. Silencing SLA expression did not affect cell viability and the insulin-producing beta-cell phenotype. Xenogeneic T-cell immune responses (p < 0.05) as well as antibody-mediated cellular-dependent immune responses (p < 0.01) were significantly decreased. Silencing SLA class I expression did not increase susceptibility to NK-cell cytotoxicity. In stirred bioreactors, tissue engineered islets showing the typical 3D-structure and morphology of ICC were assembled from SLA-silenced pancreatic cell suspensions. These data shows the feasibility to generate low immunogenic porcine ICC from transgenic pigs after single cell engineering and post-transduction islet reassembling that might serve as a robust alternative to allogeneic pancreatic islet cell transplantation.

Three-dimensional full-field vibration measurements using a handheld single-point laser Doppler vibrometer

The laser Dopper vibrometer (LDV) has become a standard instrument to perform high spatial resolution 3D vibration measurements of complex structures. An important step in the LDV measurement procedure is the estimation of the pose of the instrument relative to the test object. Existing LDV pose estimation methods rely on the use of 3D range sensors to perform this task. These sensors are costly and often their spatial resolution and accuracy is too limited for an accurate LDV pose estimation. But most importantly, existing LDV pose estimation techniques are slow. In this paper, we present a novel LDV pose estimation method based on matching of a 2D camera image and a CAD model of the test object. In contrast to existing pose estimation techniques, our method is able to track the pose of the LDV in real-time (up to about 30 LDV positions per second). This means that our method also allows LDV pose estimation when either the LDV or the object is moving. Using the real-time LDV pose tracking we present a new LDV measurement methodology where we hold instrument in our hand while moving around the test structure. This allows us to perform full-field scanning and 3D velocity measurements with a low-cost single point LDV sensor. We show that the measurement accuracy of a stationary handheld LDV measurement is about 40 dB (which is about 10 dB lower than in the case of measurements on a tripod). Furthermore, we demonstrate that the 3D vibrations measurements using the handheld vibrometer have an error of about 20% compared to classical measurements performed by combining three LDV measurements on a tripod. We illustrate our method on vibration measurements of a composite bicycle frame.

Tuning optical properties of noble metal nanoparticle-composed glasses by laser radiation

Noble metal nanoparticle composed glasses attract significant attention due to the unique optical properties that they express in the near UV and visible spectral range. These are related to the high values of the extinction cross section and nonlinear optical characteristics. In this work we study the ability of laser irradiation to induce modification of the optical properties of borosilicate glasses that contain gold nanoparticles. The process is investigated by application of laser pulses of nanosecond Nd:YAG system on glasses that consist of nanoparticles with different size and shape. The results show that at certain conditions the glass optical properties can be modified as a change of the nanoparticles plasmon resonance wavelength is observed. The influence of the laser fluence and pulse number on this effect is studied. Two fluence regimes are defined: (i) at low fluences, close to the optical properties modification threshold the increase of the laser fluence results in a blue shift of the resonance wavelength; (ii) further increase of the laser fluences induces a red shift. Similar behavior is observed by changing the number of the applied pulses. Here after application of several thousand laser pulses additional, third regime of blue shift is realized. Theoretical models based on multiparticle Mie scattering theory and heat conduction equation are applied to explain the observed modifications. On their basis and performed analyses can be concluded that the induced optical properties variations are related to modification of the nanoparticles size and shape by melting, fragmentation and coalescence. The obtained results indicate an ability of nanoparticle size and shape modifications with a high spatial resolution in 3D and can be used for fabrication of integrated optical systems.

3D printed microchannels for sub-nL NMR spectroscopy.

Nuclear magnetic resonance (NMR) experiments on subnanoliter (sub-nL) volumes are hindered by the limited sensitivity of the detector and the difficulties in positioning and holding such small samples in proximity of the detector. In this work, we report on NMR experiments on liquid and biological entities immersed in liquids having volumes down to 100 pL. These measurements are enabled by the fabrication of high spatial resolution 3D printed microfluidic structures, specifically conceived to guide and confine sub-nL samples in the sub-nL most sensitive volume of a single-chip integrated NMR probe. The microfluidic structures are fabricated using a two-photon polymerization 3D printing technique having a resolution better than 1 μm3. The high spatial resolution 3D printing approach adopted here allows to rapidly fabricate complex microfluidic structures tailored to position, hold, and feed biological samples, with a design that maximizes the NMR signals amplitude and minimizes the static magnetic field inhomogeneities. The layer separating the sample from the microcoil, crucial to exploit the volume of maximum sensitivity of the detector, has a thickness of 10 μm. To demonstrate the potential of this approach, we report NMR experiments on sub-nL intact biological entities in liquid media, specifically ova of the tardigrade Richtersius coronifer and sections of Caenorhabditis elegans nematodes. We show a sensitivity of 2.5x1013 spins/Hz1/2 on 1H nuclei at 7 T, sufficient to detect 6 pmol of 1H nuclei of endogenous compounds in active volumes down to 100 pL and in a measurement time of 3 hours. Spectral resolutions of 0.01 ppm in liquid samples and of 0.1 ppm in the investigated biological entities are also demonstrated. The obtained results may indicate a route for NMR studies at the single unit level of important biological entities having sub-nL volumes, such as living microscopic organisms and eggs of several mammalians, humans included.

3D imaging of the brain morphology and connectivity defects in a model of psychiatric disorders: MAP6-KO mice

In the central nervous system, microtubule-associated protein 6 (MAP6) is expressed at high levels and is crucial for cognitive abilities. The large spectrum of social and cognitive impairments observed in MAP6-KO mice are reminiscent of the symptoms observed in psychiatric diseases, such as schizophrenia, and respond positively to long-term treatment with antipsychotics. MAP6-KO mice have therefore been proposed to be a useful animal model for these diseases. Here, we explored the brain anatomy in MAP6-KO mice using high spatial resolution 3D MRI, including a volumetric T1w method to image brain structures, and Diffusion Tensor Imaging (DTI) for white matter fiber tractography. 3D DTI imaging of neuronal tracts was validated by comparing results to optical images of cleared brains. Changes to brain architecture included reduced volume of the cerebellum and the thalamus and altered size, integrity and spatial orientation of some neuronal tracks such as the anterior commissure, the mammillary tract, the corpus callosum, the corticospinal tract, the fasciculus retroflexus and the fornix. Our results provide information on the neuroanatomical defects behind the neurological phenotype displayed in the MAP6-KO mice model and especially highlight a severe damage of the corticospinal tract with defasciculation at the location of the pontine nuclei.

Real-Time and Multiplexed Photoacoustic Imaging of Internally Normalized Mixed-Targeted Nanoparticles.

Photoacoustic (PA) imaging is a developing diagnostic technique where multiple species can be simultaneously imaged with high spatial resolution in 3D if the absorbance spectrum of each species is distinct and separable. However, multiplexed PA imaging has been greatly limited by the availability of spectrally separable contrast agents that can be used in vivo. Toward this end, we present the formation and application of a series of poly ethylene glycol (PEG)-coated nanoparticles (NPs) with unique separable absorbance profiles suitable for simultaneous multiplexed imaging. As a proof-of-concept, we demonstrate this form of mixed-sample multiplexed imaging, using cRGD peptide surface-modified NPs with nonmodified NPs in a murine subcutaneous Lewis lung carcinoma tumor model. The simultaneous imaging of nonmodified NPs provides an "internal standard", to deconvolute the contributions of active-ligand and passive-NP targeting effects. Particles with 25% surface cRGD modification display 52 ± 22 fold higher liver to tumor ratio accumulation levels, while the same set of particles display only 9.8 ± 4 fold accumulation levels when internally normalized. The pharmacokinetic profiles of targeted and nontargeted NPs can be simultaneously tracked in real-time to study how biodistribtions of particles are affected by ligand modification. The internal normalization of control particles greatly enhances the precision and decreases the number of animals needed in studies of nanoparticle targeting. These new dyes are an enabling technology for PA imaging of NP fate and targeting. This is the first demonstration of real-time multiplexed PA imaging of mixed-targeted samples in vivo.

Saturated two-photon excitation fluorescence microscopy with core-ring illumination.

We demonstrated resolution improvement in two-photon excitation microscopy by combining saturated excitation (SAX) of fluorescence and pupil manipulation. We theoretically estimated the resolution improvement and the sidelobe effect in the point spread function with various pupil designs and found that the combination of SAX and core-ring illumination can effectively enhance the spatial resolution in 3D and suppress sidelobe artifacts. The experimental demonstration shows that the proposed technique is effective for observation with a depth of 100μm in a tissue phantom and can be applied to 3D observations of tissue samples with higher spatial resolution than conventional two-photon excitation microscopy.