A complete mathematical framework for preclinical optical imaging (OI) support comprising bioluminescence imaging (BLI), fluorescence surface imaging (FSI) and fluorescence optical tomography (FOT) is presented in which optical data is acquired by means of a microlens array (MLA) based light detector (MLA-D). The MLA-D has been developed to enable unique OI, especially in synchromodal operation with secondary imaging modalities (SIM) such as positron emission tomography (PET) or magnetic resonance imaging (MRI). An MLA-D consists of a (large-area) photon sensor array, a matched MLA for field-of-view definition, and a septum mask of specific geometry made of anodized aluminum that is positioned between the sensor and the MLA to suppresses light cross-talk and to shield the sensor's radiofrequency interference signal (essential when used inside an MRI system). The software framework, while freely parameterizable for any MLA-D, is tailored towards an OI prototype system for preclinical SIM application comprising a multitude of cylindrically assembled, gantry-mounted, simultaneously operating MLA-D's. Besides the MLA-D specificity, the framework incorporates excitation and illumination light-source declarations of large-field and point geometry to facilitate multispectral FSI and FOT as well as three-dimensional object recognition. When used in synchromodal operation, reconstructed tomographic SIM volume data can be used for co-modal image fusion and also as a prior for estimating the imaged object's 3D surface by means of gradient vector flow. Superimposed planar (without object prior) or surface-aligned inverse mapping can be performed to estimate and to fuse the emission light map with the boundary of the imaged object. Triangulation and subsequent optical reconstruction (FOT) or constrained flow estimation (BLI), both including the possibility of SIM priors, can be performed to estimate the internal three-dimensional emission light distribution. The framework is susceptible to a number of variables controlling convergence and computational speed. Utilization and performance is illustrated on experimentally acquired data employing the OI prototype system in stand-alone operation, and when integrated into an unmodified preclinical PET system performing synchromodal BLI-PET in vivo imaging.

An established challenge with fluorescence molecular tomography is inconsistent accuracy in quantifying recovered fluorescent contrast with changes in target depth. This work examines the idea that through optimization of the source and detector placement more consistent contrast could be recovered regardless of imaging depth. A simulation study was performed to examine distributions of optical projection measurements result in a uniform summed sensitivity function, showing that half-reflectance and transmission measurements are the most important for accurate and consistent target recovery as a function of depth. As a corollary to the importance of uniform summed sensitivity, translation modulated reconstructions are shown to provide improvements in fluorescence tomography.

We introduce an entirely new technique, termed Photo-Magnetic Imaging (PMI), which overcomes the limitation of pure optical imaging and provides optical absorption at MRI spatial resolution. PMI uses laser light to heat the medium under investigation and employs MR thermometry for the determination of spatially resolved optical absorption in the probed medium. A FEM-based PMI forward solver has been developed by modeling photon migration and heat diffusion in tissue to compare simulation results with measured MRI maps. We have successfully performed PMI using 2.5 cm diameter agar phantom with two low optical absorption contrast (x 4) inclusions under the ANSI limit. Currently, we are developing the PMI inverse solver and undertaking further phantom and in vivo experiments.

Low spatial resolution due to strong tissue scattering is one of the main barriers that prevent the wide-spread use of fluorescence tomography. To overcome this limitation, we previously demonstrated a new technique, temperature modulated fluorescence tomography (TM-FT), which relies on key elements: temperature sensitive ICG loaded pluronic nanocapsules and high intensity focused ultrasound (HIFU), to combine the sensitivity of fluorescence imaging with focused ultrasound resolution. While conventional fluorescence tomography measurements are acquired, the tissue is scanned by a HIFU beam and irradiated to produce a local hot spot, in which the temperature increases nearly 5K. The fluorescence emission signal measured by the optical detectors varies drastically when the hot spot overlays onto the location of the temperature dependent nanocapsules. The small size of the focal spot (~1.4 mm) up to a depth of 6 cm, allows imaging the distribution of these temperature sensitive agents with not only high spatial resolution but also high quantitative accuracy in deep tissue using a proper image reconstruction algorithm. Previously we have demonstrated this technique with a phantom study with nanocapsules sensitive to 20-25°C range. In this work, we will show the first nanocapsules optimized for in vivo animal imaging.

Over 70% of military casualties resulting from the current conflicts sustain major extremity injuries. Of these the majority are caused by blasts from improvised explosive devices. The resulting injuries include traumatic amputations, open fractures, crush injuries, and acute vascular disruption. Critical tissue ischemia—the point at which ischemic tissues lose the capacity to recover—is therefore a major concern, as lack of blood flow to tissues rapidly leads to tissue deoxygenation and necrosis. If left undetected or unaddressed, a potentially salvageable limb may require more extensive debridement or, more commonly, amputation. Predicting wound outcome during the initial management of blast wounds remains a significant challenge, as wounds continue to “evolve” during the debridement process and our ability to assess wound viability remains subjectively based. Better means of identifying critical ischemia are needed. We developed a swine limb ischemia model in which two imaging modalities were combined to produce an objective and quantitative assessment of wound perfusion and tissue viability. By using 3 Charge-Coupled Device (3CCD) and Infrared (IR) cameras, both surface tissue oxygenation as well as overall limb perfusion could be depicted. We observed a change in mean 3CCD and IR values at peak ischemia and during reperfusion correlate well with clinically observed indicators for limb function and vitality. After correcting for baseline mean R-B values, the 3CCD values correlate with surface tissue oxygenation and the IR values with changes in perfusion. This study aims to not only increase fundamental understanding of the processes involved with limb ischemia and reperfusion, but also to develop tools to monitor overall limb perfusion and tissue oxygenation in a clinical setting. A rapid and objective diagnostic for extent of ischemic damage and overall limb viability could provide surgeons with a more accurate indication of tissue viability. This may help reducing the number of surgical interventions required, by aiding surgeons in identifying and demarcating areas of critical tissue ischemia, so that a more adequate debridement may be performed. This would have obvious benefits of reducing patient distress and decreasing both the overall recovery time and cost of rehabilitation.

Elasticity is an important indicator of tissue health, with increased stiffness pointing to an increased risk of cancer. We investigated a tissue elasticity measurement method using forward and inversion algorithms for the application of early breast tumor identification. An optical based elasticity measurement system is developed to capture images of the embedded lesions using total internal reflection principle. From elasticity images, we developed a novel method to estimate the elasticity of the embedded lesion using 3-D finite-element-model-based forward algorithm, and neural-network-based inversion algorithm. The experimental results showed that the proposed characterization method can be diffierentiate the benign and malignant breast lesions.

Clinical interventions can cause changes in tissue perfusion, oxygenation or temperature. Real-time imaging of these phenomena could be useful for surgical strategy or understanding of physiological regulation mechanisms. Two noncontact imaging techniques were applied for imaging of large tissue areas: LED based multispectral imaging (MSI, 17 different wavelengths 370 nm-880 nm) and thermal imaging (7.5 to 13.5 μm). Oxygenation concentration changes were calculated using different analyzing methods. The advantages of these methods are presented for stationary and dynamic applications. Concentration calculations of chromophores in tissue require right choices of wavelengths The effects of different wavelength choices for hemoglobin concentration calculations were studied in laboratory conditions and consequently applied in clinical studies. Corrections for interferences during the clinical registrations (ambient light fluctuations, tissue movements) were performed. The wavelength dependency of the algorithms were studied and wavelength sets with the best results will be presented. The multispectral and thermal imaging systems were applied during clinical intervention studies: reperfusion of tissue flap transplantation (ENT), effectiveness of local anesthetic block and during open brain surgery in patients with epileptic seizures. The LED multispectral imaging system successfully imaged the perfusion and oxygenation changes during clinical interventions. The thermal images show local heat distributions over tissue areas as a result of changes in tissue perfusion. Multispectral imaging and thermal imaging provide complementary information and are promising techniques for real-time diagnostics of physiological processes in medicine.

We have previously demonstrated the utilization of spatially co-registered diffuse optical tomography (DOT) and digital breast tomosynthesis (DBT) for joint breast cancer diagnosis. However, clinical implementation of such a multi-modality approach may require development of integrated DOT/DBT imaging scanners, which can be costly and time-consuming. Exploring effective image registration methods that combine the diagnostic information from a standalone DOT measurement and a separate mammogram can be a cost-effective solution, which may eventually enable adding functional optical assessment to all previously installed digital mammography systems. In this study, we investigate a contour-based image registration method to convert independent optical and x-ray scans into co-registered datasets that can benefit from a joint image analysis. The breast surface used in 3D optical DOT reconstruction is registered with the breast contour line extracted from an x-ray mammogram acquired separately. This allows us to map the 2D mammogram to the optical measurement space and build structural constraints for optical image reconstruction. A non-linear reconstruction utilizing structure-priors is then performed to produce hemoglobin maps with improved resolution. To validate this approach, we used a set of tumor patient measurements with simultaneous DOT/DBT and separate 2D mammographic scans. The images recovered from the registration procedure derived from DOT and 2D mammogram present similar image quality compared to those recovered from co-registered DOT/DBT measurements.

There is a pressing clinical need to provide image guidance during surgery. Currently, assessment of tissue that needs to be resected or avoided is performed subjectively, leading to a large number of failures, patient morbidity, and increased healthcare costs. Because near-infrared (NIR) optical imaging is safe, noncontact, inexpensive, and can provide relatively deep information (several mm), it offers unparalleled capabilities for providing image guidance during surgery. These capabilities are well illustrated through the clinical translation of fluorescence imaging during oncologic surgery. In this work, we introduce a novel imaging platform that combines two complementary NIR optical modalities: oxygenation imaging and fluorescence imaging. We validated this platform during facial reconstructive surgery on large animals approaching the size of humans. We demonstrate that NIR fluorescence imaging provides identification of perforator arteries, assesses arterial perfusion, and can detect thrombosis, while oxygenation imaging permits the passive monitoring of tissue vital status, as well as the detection and origin of vascular compromise simultaneously. Together, the two methods provide a comprehensive approach to identifying problems and intervening in real time during surgery before irreparable damage occurs. Taken together, this novel platform provides fully integrated and clinically friendly endogenous and exogenous NIR optical imaging for improved image-guided intervention during surgery.

A compact prototype device for diagnostic imaging of skin has been developed and tested. Polarized LED light at several spectral regions is used for illumination, and round skin spot of diameter 30mm is imaged by a CMOS sensor via crossoriented polarizing filter. Four consecutive imaging series are performed: (1) RGB image at white LED illumination for revealing subcutaneous structures; (2) four spectral images at narrowband LED illumination (450nm, 540nm, 660nm, 940nm) for mapping of the main skin chromophores; (3) video-imaging under green LED illumination for mapping of skin blood perfusion; (4) autofluorescence video-imaging under UV (365nm) LED irradiation for mapping of the skin fluorophores. Design details of the device as well as preliminary results of clinical tests are presented.

Since diffuse optical tomography (DOT) is a low spatial resolution modality, it is desirable to validate its quantitative accuracy with another well-established imaging modality, such as magnetic resonance imaging (MRI). In this work, we have used a polymer based bi-functional MRI-optical contrast agent (Gd-DTPA-polylysine-IR800) in collaboration with GE Global Research. This multi-modality contrast agent provided not only co-localization but also the same kinetics, to cross-validate two imaging modalities. Bi-functional agents are injected to the rats and pharmacokinetics at the bladder are recovered using both optical and MR imaging. DOT results are validated using MRI results as "gold standard"

Upregulate levels of expression and activity of membrane H⁺ ion pumps in cancer cells drives the extracellular pH (pH_e,) to values lower than normal. Furthermore, disregulated pH is indicative of the changes in glycolytic metabolism in tumor cells and has been shown to facilitate extracellular tissue remodeling during metastasis Therefore, measurement of pH_e could be a useful cancer biomarker for diagnostic and therapy monitoring evaluation. Multimodality in-vivo imaging of pH_e in tumorous tissue in a mouse dorsal skin fold window chamber (DSFWC) model is described. A custom-made plastic window chamber structure was developed that is compatible with both imaging optical and MR imaging modalities and provides a model system for continuous study of the same tissue microenvironment on multiple imaging platforms over a 3-week period. For optical imaging of pH_e, SNARF-1 carboxylic acid is injected intravenously into a SCID mouse with an implanted tumor. A ratiometric measurement of the fluorescence signal captured on a confocal microscope reveals the pH_e of the tissue visible within the window chamber. This imaging method was used in a preliminary study to evaluate sodium bicarbonate as a potential drug treatment to reverse tissue acidosis. For MR imaging of pH_e the chemical exchange saturation transfer (CEST) was used as an alternative way of measuring pHe in a DSFWC model. ULTRAVIST®, a FDA approved x-ray/CT contrast agent has been shown to have a CEST effect that is pH dependent. A ratiometric analysis of water saturation at 5.6 and 4.2 ppm chemical shift provides a means to estimate the local pH_e.

Multimodal imaging is quickly becoming a standard in pre-clinical studies, and new developments have already confirmed the strength of acquiring and analyzing parallel information obtained in a single imaging session. One such application is the introduction of an internal reference moiety (e.g. radioisotope) to an activatable fluorescent probe. One of the limitations of this approach consists of working at concentrations which are within the overlapping range of sensitivities of each modality. In the case of PET/Fluorescence imaging, this range is in the order of 10^-9 nM. Working in epi-illumination fluorescence imaging at such concentrations remains challenging. Here, we present in vitro and in vivo detection limits of a new fluorescent compound.

A fluorescence optical tomography approach that extends time reversal optical tomography (TROT) to locate fluorescent targets embedded in a turbid medium is introduced. It uses a multi-source illumination and multi-detector signal acquisition scheme, along with TR matrix formalism, and multiple signal classification (MUSIC) to construct pseudo-image of the targets. The samples consisted of a single or two small tubes filled with water solution of Indocyanine Green (ICG) dye as targets embedded in a 250 mm × 250 mm × 60 mm rectangular cell filled with Intralipid-20% suspension as the scattering medium. The ICG concentration was 1μM, and the Intralipid-20% concentration was adjusted to provide ~ 1-mm transport length for both excitation wavelength of 790 nm and fluorescence wavelength around 825 nm. The data matrix was constructed using the diffusely transmitted fluorescence signals for all scan positions, and the TR matrix was constructed by multiplying data matrix with its transpose. A pseudo spectrum was calculated using the signal subspace of the TR matrix. Tomographic images were generated using the pseudo spectrum. The peaks in the pseudo images provided locations of the target(s) with sub-millimeter accuracy. Concurrent transmission TROT measurements corroborated fluorescence-TROT findings. The results demonstrate that TROT is a fast approach that can be used to obtain accurate three-dimensional position information of fluorescence targets embedded deep inside a highly scattering medium, such as, a contrast-enhanced tumor in a human breast.

At present, the most widely accepted forward model in diffuse optical tomography (DOT) is the diffusion equation, which is derived from the radiative transfer equation by employing the P1 approximation. However, due to its validity restricted to highly scattering regions, this model has several limitations for the whole-body imaging of small-animals, where some cavity and low scattering areas exist. To overcome the difficulty, we presented a Graphic-Processing- Unit(GPU) implementation of Monte-Carlo (MC) modeling for photon migration in arbitrarily heterogeneous turbid medium, and, based on this GPU-accelerated MC forward calculation, developed a fast, universal DOT image reconstruction algorithm. We experimentally validated the proposed method using a continuous-wave DOT system in the photon-counting mode and a cylindrical phantom with a cavity inclusion.

To cope with the low quantification in the established optical topography that originates from the excessively simplified computation model based on the modified Lambert-Beer’s Law (MLBL), we propose a least-squares fitting scheme for time-domain optical topography that seeks for data matching between the time-resolved measurement and the model prediction calculated by analytically solving the time-domain diffusion equation in semi-infinite geometry. Our simulative and phantom experiments demonstrate that the proposed curve-fitting method is overall superior to the conventional MLBL-based one in quantitative performance.

Color intensity projections (CIPs) has been employed to improve the accuracy and reduce the workload of interpreting a series of grayscale images by summarizing the grayscale images in a single color image. CIPs – which has been applied to grayscale images in angiography, 4D CT, nuclear medicine and astronomy – uses the hue, saturation and brightness of the color image to encode the summary information. In CIPs, when a pixel has the same value over the grayscale images, the corresponding pixel in the color image has the identical grayscale color. The arrival time of a signal at each pixel, such as the arrival time of contrast in angiography, is often encoded in the hue (red-yellow-green-light blue-blue-purple) of the corresponding pixel in the color image. In addition, the saturation and brightness of each pixel in the color image encodes the amplitude range and amplitude maximum of the corresponding pixel in the grayscale images. In previous applications of CIPs the hue has been limited to less than one cycle over the color image to avoid the aliasing due to a hue corresponding to more than one arrival time. However, sometimes in applications such as angiography and astronomy, in some instances the aliasing due to increasing the number of cycles of hue over the color image is tolerable as it increases the resolution of arrival time. Key to applying hue cycling effectively is interpolating several grayscale images between each pair of grayscale images. Ideally, the interpreter is allowed to adjust the amount of hue cycling in realtime to find the best setting for each particular CIPs image. CIPs with hue cycling should be a valuable tool in many fields where interpreting a series of grayscale images is required.

Changes in body temperature have clinically significant effects on nerve excitation. However, the mechanisms of these effects still further explored. In this paper, we studied the effects of temperature on the propagation properties of action potential on unmyelinated afferents by action-potential-sensitive second harmonic generation (SHG). A mathematical model was presented which combining with SHG imaging to study the action potential properties in an unmylinated fiber affected by temperature. The results showed that the action potential propagation along an unmyelinated fiber was sensitively probed by optical SHG imaging in real time. Meanwhile, the action potential properties on an unmyelinated fiber is obviously changed at innocuous temperatures through analyzing the changes of action-potential-sensitive SHG signals, including the increase of conduction velocity and the decrease of duration at higher temperatures. Our study indicates that optical second harmonic generation imaging may be a potential tool to investigate the underlying mechanisms of complex physiological phenomena such as nerve excitation.