This PDF file contains the front matter associated with SPIE Proceedings Volume 7567, including the Title Page, Copyright information, Table of Contents, Introduction, and the Conference Committee listing.

Validation of imaging technologies is becoming increasingly important as imaging begins to take a role as a biomarker. Activities such as drug development, guided intervention, patient stratification, and therapy response assessment rely on very high measurements of sensitivity and specificity at the earliest possible time point. Imaging is capable of providing those measurements, but without validation of the results, clinical acceptance will be delayed. It must be pointed out, however, that validation is different from verification, and the role of phantoms is different for these two operations. This paper discusses the need for multiple phantoms in the role of validation in order to present optical imaging devices with the extremes of tissue states expected in real-world operation.

Ongoing multi-center clinical trials are crucial for Biophotonics to gain acceptance in medical imaging. In these trials, quality control (QC) and assurance (QA) are key to success and provide "data insurance". Quality control and assurance deal with standardization, validation, and compliance of procedures, materials and instrumentation. Specifically, QC/QA involves systematic assessment of testing materials, instrumentation performance, standard operating procedures, data logging, analysis, and reporting. QC and QA are important for FDA accreditation and acceptance by the clinical community. Our Biophotonics research in the Network for Translational Research in Optical Imaging (NTROI) program for breast cancer characterization focuses on QA/QC issues primarily related to the broadband Diffuse Optical Spectroscopy and Imaging (DOS/I) instrumentation, because this is an emerging technology with limited standardized QC/QA in place. In the multi-center trial environment, we implement QA/QC procedures: 1. Standardize and validate calibration standards and procedures. (DOS/I technology requires both frequency domain and spectral calibration procedures using tissue simulating phantoms and reflectance standards, respectively.) 2. Standardize and validate data acquisition, processing and visualization (optimize instrument software-EZDOS; centralize data processing) 3. Monitor, catalog and maintain instrument performance (document performance; modularize maintenance; integrate new technology) 4. Standardize and coordinate trial data entry (from individual sites) into centralized database 5. Monitor, audit and communicate all research procedures (database, teleconferences, training sessions) between participants ensuring "calibration". This manuscript describes our ongoing efforts, successes and challenges implementing these strategies.

In this work we present contrast measurement phantoms for optical coherence tomography. In this initial study, contrast has been evaluated from OCT images of a USAF-1951 bar target, buried beneath scattering layers of different density. Preliminary results indicate that scattering does not degrade imaging contrast significantly, however further work is required to verify these findings. This work has important implications from a quality control perspective as well for OCT inter-comparisons at, for example, different wavebands.

Microscopes are being developed for use in living animals, and even humans, to image microanatomical changes and molecular markers that are associated with disease. Phantoms that can be used to evaluate the performance characteristics of these systems have not been well described or standardized. We have been developing the tools to evaluate a dual-axis confocal (DAC) microscope design to optimize the features required for in vivo diagnosic imaging, and these may have features that are useful for evaluation of other such devices. We have performed diffraction-theory modeling, Monte-Carlo scattering simulations, reflectance experiments in tissue phantoms, and tissue-imaging validations. First, we determined how scattering from tissue deteriorates the diffraction-limited transverse and vertical responses in reflectance DAC imaging. Specifically, the vertical and transverse responses of the DAC to a plane reflector and a knife edge, respectively, were measured at various depths in an Intralipid scattering phantom. Comparisons were made with both diffraction-theory and Monte-Carlo scattering simulations. Secondly, as a practical demonstration of deep-tissue fluorescence microscopy, three-dimensional fluorescence images were obtained in thick human biopsy samples. These results demonstrate that the efficient rejection of scattered light in a DAC microscope enables deep optical sectioning in tissue. Finally, we will discuss our needs and plans for similar tissue-phantom experiments to validate the performance of multimodal optical- and ultrasound-imaging platforms under development. As devices are developed for the imaging of epithelial surfaces and substructures, standardized phantoms that represent the multilayered anatomical features of these tissues will need to be developed.

With the development of multilayer models for the analysis of quantitative spectroscopic techniques, there is a need to generate controlled and stable phantoms capable of validating these new models specific to the particular instrument performance and/or probe geometry. Direct applications for these multilayer phantoms include characterization or validation of depth penetration for specific probe geometries or describing layer specific sensitivity of optical instrumentation. We will present a method of producing interchangeable silicone phantoms that vary in thickness from 90 microns up to several millimeters which can be combined to produce multilayered structures to mimic optical properties of physiologic tissues such as skin. The optical properties of these phantoms are verified through inverse addingdoubling methods and the homogeneous distribution of optical properties will be discussed.

Glioblastoma are brain tumors currently incurable, however, optimized treatment gives better prognosis and quality of life. In case of surgical treatment, there is still need to help surgeons to determine whether a tissue is tumorous or not. Within the framework of the design of a new autofluorescence probe for this issue, optically calibrated gel phantoms have been developed using "tumorous" inclusions in a "healthy" environment. Depending on "tumor" shape, size and localization, the sensitivity of the probe is evaluated. The probe sensitivity for fluorescence spectroscopy will be presented. The probe configuration is also taken into account and compared to simulated results.

MCML1.2.2-2000 code was used to simulate light distribution in Lipovenos^R 10% (Lp) layers with various thicknesses illuminated by red laser. Light fluence distribution at the layer bottom and fluence profile along a plane distant 5.5 mm from the laser beam were calculated. The results show that the light transmitted to the bottom of the sample has a Gaussian distribution with widths that increase linearly with the thickness. Also, the maximum light intensity and the total fluence transmitted across the sample have exponential decay behavior with thickness. An experiment has been carried out, acquiring, with a CCD camera, pictures of light transmitted and scattered at 90° from a cuvette containing different quantities of Lp, illuminated from the top with He-Ne laser. The experimental results show that the maximum intensity of transmitted light has an asymptotic exponential behavior with the sample thickness, very similar to the simulation. Gaussian curves fitted to the experimental results have widths similar to the simulated ones. The simulated light profile at 5.5 mm from the incidence plane is very similar to the variation of scattered light intensity with depth. We conclude that images of illuminated tissue combined with MCS can contribute with evaluation of light distribution inside tissue.

Solid tissue phantom are the preferred tool for the development, validation, testing and calibration of photon migration instrument. Accuracy, or trueness, of the optical properties of reference phantoms is of the utmost importance as they will be used as the conventional true value against which instrument errors will be evaluated. A detailed quantitative analysis of the uncertainty of time-resolved transmittance characterization of solid optical tissue phantom is presented. Random error sources taken into account are Poisson noise of the photon counting process, additive dark count noise and instrument response function stability. Systematic error sources taken into account are: phantom thickness uncertainty, refractive index uncertainty, time correlated single photon counting system time base calibration uncertainty. Correction procedures for these systematic errors are presented whenever a correction is possible.

We fabricated permanent solid polyurethane-based phantoms in which fluorophores were homogeneously incorporated. For this study, fluorophores of three different families were used: Cyanines, Alexa Fluor and Quantum Dots. The goal of this study was to evaluate the impact of casting the fluorophores in a polyurethane matrix on their optical properties, more specifically the absorbance, molecular extinction coefficient, emission of fluorescence and the resultant fluorescence intensity. All measurements were carried out with 5 concentrations of each fluorophores embedded in polyurethane and in solution. Stability over time was also monitored for a three months period. The casting of fluorophores affects the optical properties of the three dyes under study. The max absorbance, the fluorescence emission and intensity along with the molar extinction coefficient were all affected. Quantum dots behave differently to the cyanine and Alexa Fluor dyes. It was also observed that the incorporation of dyes enables long-term stability of the fluorescence signal.

Laser light propagation in soft tissues is important because of the growing biomedical applications of lasers and the need to optically characterize the biological media. Following previous developments of the group, we have developed low cost models, Phantoms, of soft tissue. The process was developed in a clean room to avoid the medium contamination. Each model was characterized by measuring the refractive index, and spectral reflectance and transmittance. To study the laser light propagation, each model was illuminated with a clean beam of laser light, using sources such as He-Ne (632nm) and DPSSL (473 nm). Laterally scattered light was imaged and these images were digitally processed. We analyzed the intensity distribution of the scattered radiation in order to obtain details of the beam evolution in the medium. Line profiles taken from the intensity distribution surface allow measuring the beam spread, and to find expressions for the longitudinal (along the beam incident direction) and transversal (across the beam incident direction) intensities distributions. From these behaviors, the radiation penetration depth and the total coefficient of extinction have been determined. The multiple scattering effects were remarkable, especially for the low wavelength laser beam.

Phantoms with controlled optical properties are often used for calibration and standardization. The phantoms are typically prepared by adding absorbers and scatterers to a clear host material. It is usually assumed that the scatterers and absorbers are uniformly dispersed within the medium. To explore the effects of this assumption, we prepared paired sets of polyurethane phantoms (both with identical masses of absorber, India ink and scatterer, titanium dioxide). Polyurethane phantoms were made by mixing two polyurethane parts (a and b) together and letting them cure in a polypropylene container. The mixture was degassed before curing to ensure a sample without bubbles. The optical properties were controlled by mixing titanium dioxide or India ink into polyurethane part (a or b) before blending the parts together. By changing the mixing sequence, we could change the aggregation of the scattering and absorbing particles. Each set had one sample with homogeneously dispersed scatterers and absorbers, and a second sample with slightly aggregated scatterers or absorbers. We found that the measured transmittance could easily vary by a factor of twenty. The estimated optical properties (using the inverse adding-doubling method) indicate that when aggregation is present, the optical properties are no longer proportional to the concentrations of absorbers or scatterers.

The initial steps in fabricating a multimodality imaging phantom for combined diffuse optical tomography and ultrasound tomography (DOT-UST) are completed. Phantoms are intended to mimic the optical and acoustic properties of breast tissue for near infrared light and ultrasound in the vicinity of 2 MHz. So far, a prototype ultrasound tomography system has been designed and the acoustic attenuation coefficient of glass beads has been characterized. Furthermore, 8 cm diameter homogeneous cylindrical phantoms have been successfully constructed and it has been shown that an inclusion with object to background contrast of three can be comfortably detected with the prototype system.

We report preliminary results toward making artery phantoms for Optical Coherence Tomography (OCT) that also exhibit mechanical properties similar to arteries for large deformation regimes. A matrix of PVA cryogels is used to obtain the strain hardening effect characteristic of arteries. Means of adjusting the optical properties of PVA cryogels are investigated and the resulting mechanical properties are characterized.

We describe an optical tissue phantom that enables the simulation of drug extravasation from microvessels and validates computational compartmental models of drug delivery. The phantom consists of a microdialysis tubing bundle to simulate the permeable blood vessels, immersed in either an aqueous suspension of titanium dioxide (TiO2) or a TiO2 mixed agarose scattering medium. Drug administration is represented by a dye circulated through this porous microdialysis tubing bundle. Optical pharmacokinetic (OP) methods are used to measure changes in the absorption coefficient of the scattering medium due to the arrival and diffusion of the dye. We have established particle sizedependent concentration profiles over time of phantom drug delivery by intravenous (IV) and intra-arterial (IA) routes. Additionally, pharmacokinetic compartmental models are implemented in computer simulations for the conditions studied within the phantom. The simulated concentration-time profiles agree well with measurements from the phantom. The results are encouraging for future optical pharmacokinetic method development, both physical and computational, to understand drug extravasation under various physiological conditions.

Many advantages of biomedical optical imaging modalities include low cost, portability, no radiation hazard, molecular sensitivity, and real-time non-invasive measurements of multiple tissue parameters. However, clinical acceptance of optical imaging is hampered by the lack of calibration standards and validation techniques. In this context, developing phantoms that simulate tissue structural, functional, and molecular properties is important for reliable performance and successful translation of biomedical optical imaging techniques to clinical applications. Over the years, we have developed various tissue simulating phantoms to validate imaging algorithms, to optimize instrument performance, to test contrast agents, and to calibrate acquisition systems. We also developed phantoms with multimodal contrasts for co-registration between different imaging modalities. In order to study tissue dynamic changes during medical intervention, we develop gel wax phantoms to simulate tissue optical and mechanical dynamics in response to compression load. We also dispersed heat sensitive microbubbles in agar agar gel phantoms to simulate heatinduced tissue coagulative necrosis in a cancer ablation procedure. The phantom systems developed in our lab have the potential to provide standardized traceable tools for multimodal imaging and image-guided intervention.

Background: Clinical ultrasound (US) uses ultrasonic scattering contrast to characterize subcutaneous anatomic structures. Photoacoustic (PA) imaging detects the functional properties of thick biological tissue with high optical contrast. In the case of image-guided cancer ablation therapy, simultaneous US and PA imaging can be useful for intraoperative assessment of tumor boundaries and ablation margins. In this regard, accurate co-registration between imaging modalities and high sensitivity to cancer cells are important. Methods: We synthesized poly-lactic-co-glycolic acid (PLGA) microbubbles (MBs) and nanobubbles (NBs) encapsulating India ink or indocyanine green (ICG). Multiple tumor simulators were fabricated by entrapping ink MBs or NBs at various concentrations in gelatin phantoms for simultaneous US and PA imaging. MBs and NBs were also conjugated with CC49 antibody to target TAG-72, a human glycoprotein complex expressed in many epithelial-derived cancers. Results: Accurate co-registration and intensity correlation were observed in US and PA images of MB and NB tumor simulators. MBs and NBs conjugating with CC49 effectively bound with over-expressed TAG-72 in LS174T colon cancer cell cultures. ICG was also encapsulated in MBs and NBs for the potential to integrate US, PA, and fluorescence imaging. Conclusions: Multifunctional MBs and NBs can be potentially used as a general contrast agent for multimodal intraoperative imaging of tumor boundaries and therapeutic margins.