Time-correlated single-photon counting (TCSPC) allows to achieve picosecond-precision measurements for low-light signals. However, TCSPC suffers from pile-up distortion, constraining the acquisition rate to 1-5% of the laser rate. To overcome the issue, our research focuses on high-rate TCSPC methodologies: in 2017 we reported a hardware acquisition approach, that has been translated into a real system, guaranteeing low distortion at 32 Mcps. This talk provides an overview on the research project, and in particular on the two validation campaigns carried out in fluorescence and lidar measurements, and on our first on-field experiment, i.e. the application of the technique to a single-pixel camera.
Multispectral fluorescence lifetime imaging microscopy (λFLIM) is a powerful optical technique to investigate biological processes, which generally requires long acquisition time. Single Pixel Camera (SPC) is an imaging architecture base on Compressive Sensing (CS) techniques which allows to strongly reduce the acquisition time while preserving the information content at the cost of an increased computational time. In this work we present a λFLIM microscope based on CS-SPC architecture. We have tested the multiscale capability of the system by merging SPC zooming with data fusion and proposed a fast fitting framework, which runs in parallel with the acquisition, allowing a fast visualization.
We present initial evidence of the SOLUS potential for the multimodal non-invasive diagnosis of breast cancer by describing the correlation between optical and standard radiological data and analyzing a case study.
Multispectral fluorescence lifetime microscopy (FLIM) is a valuable tool for biomedical and environmental applications. A multidimensional acquisition scheme (space, time, spectrum) provides high information content and the drawback of long acquisition/processing times. Compressive Sensing (CS) combined with Single-Pixel Camera (SPC) acquisition scheme has been proposed as a strategy to reduce the number of measurements. We present a multispectral FLIM system based on SPC, CS and data fusion (DF) with a high-resolution camera to strongly reduce the acquisition time. We adopted a novel method for TCSPC to increase the count-rate. The system is characterized and validated on a cellular sample.
Time-resolved multispectral fluorescence microscopy provides a 4D hypercube dataset with high specificity for cellular examination. However, this is generally obtained by significantly increasing the measurement time, which is quite limiting for in vivo measurement or with photosensitive sample. It is possible to reduce the measurement effort with a novel microscopy framework exploiting compressive sensing based on single-pixel camera. In this work, we present a compressive sensing system and validate it with a cellular sample. Data fusion with a high-resolution camera image allows us to tackle the well-known problem of low-resolution in single-pixel imaging.
SignificanceInterstitial fiber-based spectroscopy is gaining interest for real-time in vivo optical biopsies, endoscopic interventions, and local monitoring of therapy. Different from other photonics approaches, time-domain diffuse optical spectroscopy (TD-DOS) can probe the tissue at a few cm distance from the fiber tip and disentangle absorption from the scattering properties. Nevertheless, the signal detected at a short distance from the source is strongly dominated by the photons arriving early at the detector, thus hampering the possibility of resolving late photons, which are rich in information about depth and absorption.AimTo fully benefit from the null-distance approach, a detector with an extremely high dynamic range is required to effectively collect the late photons; the goal of our paper is to test its feasibility to perform TD-DOS measurements at null source–detector separations (NSDS).ApproachIn particular, we demonstrate the use of a superconducting nanowire single photon detector (SNSPD) to perform TD-DOS at almost NSDS ( ≈ 150 μm ) by exploiting the high dynamic range and temporal resolution of the SNSPD to extract late arriving, deep-traveling photons from the burst of early photons.ResultsThis approach was demonstrated both on Monte Carlo simulations and on phantom measurements, achieving an accuracy in the retrieval of the water spectrum of better than 15%, spanning almost two decades of absorption change in the 700- to 1100-nm range. Additionally, we show that, for interstitial measurements at null source–detector distance, the scattering coefficient has a negligible effect on late photons, easing the retrieval of the absorption coefficient.ConclusionsUtilizing the SNSPD, broadband TD-DOS measurements were performed to successfully retrieve the absorption spectra of the liquid phantoms. Although the SNSPD has certain drawbacks for use in a clinical system, it is an emerging field with research progressing rapidly, and this makes the SNSPD a viable option and a good solution for future research in needle guided time-domain interstitial fiber spectroscopy.
We demonstrate a novel realization of Interstitial fiber, broadband, Time Domain Diffuse Optical Spectroscopy (TD-DOS) in Null Source-Detector separation (NSDS) approach without temporal gating, by using a Super-conducting Nanowire single photon detector (SNSPD) for acquisition. We test its feasibility by performing Monte Carlo simulations and comparing the absorption retrieval of the SNSPD with an ideal scenario and a standard Silicon Photomultiplier (SiPM). Consequently, as per the MEDPHOT protocol, we test experimentally, the absorption linearity of the system on tissue-equivalent liquid phantoms and demonstrate the scattering independent retrieval of the absorption spectrum of water using Intralipid phantoms in the wavelength range of 600-1100 nm.
By exploiting the recent components miniaturization trend, we realize a small and cheap multifunctional time-resolved (TR) single-photon detection chain. It is based on 16 channels, which can be configured either as 16 independently located channels for TR diffuse optical tomography or as a linear array for parallel TR fluorescence spectroscopy. Both applications require a detector with high time resolution and high light harvesting capability (i.e., large active area and detection efficiency). Thus, each detection channel contains a 1.3 x 1.3 mm2 active area silicon photomultiplier and its home-made electronics specifically designed for avalanche sensing and amplification, capable of optimizing the single-photon timing resolution despite the miniaturization. In this study we describe the timing performances of a first 8- channel prototype and its first application in fluorescence lifetime sensing. Then, we show the capability of the whole 16-channel array in detecting absorption changes within a homogeneous scattering medium. We have been able to obtain a single-photon timing resolution of almost 60 ps, that is close to the best ever achieved with this kind of detector. For the validation in fluorescence lifetime sensing, the fluorescence signal acquired by the proposed prototype is comparable to the one obtained using a state-of-the-art setup based on a PMT detector. In the validation in diffuse optics, we clearly detected the absorption perturbation. This confirms the suitability of this stackable solid-state detector array for both applications.
A machine learning classification algorithm is applied to the SOLUS database to discriminate benign and malignant breast lesions, based on absorption and composition properties retrieved through diffuse optical tomography. The Mann-Whitney test indicates oxy-hemoglobin (p-value = 0.0007) and lipids (0.0387) as the most significant constituents for lesion classification, but work is in progress for further analysis. Together with sensitivity (91%), specificity (75%) and the Area Under the ROC Curve (0.83), special metrics for imbalanced datasets (27% of malignant lesions) are applied to the machine learning outcome: balanced accuracy (83%) and Matthews Correlation Coefficient (0.65). The initial results underline the promising informative content of optical data.
Time-Correlated Single Photon Counting (TCSPC) represents a fundamental tool for the investigation of biological light signals. Unfortunately, due to pile-up distortion the photon acquisition rate must be kept below few percents of the laser rate, thus increasing the acquisition time.
Recently, we proposed a single-channel TCSPC system allowing us to overcome pile-up by matching the detector dead time to the laser period. In this work, we perform on-field fluorescence measurements with this system showing that an acquisition speed of 32 Mcps can be reached without significant distortion. Thanks to the promising results, we are now developing a multi-channel module based on the same acquisition technique.
Significance: Multi-laboratory initiatives are essential in performance assessment and standardization—crucial for bringing biophotonics to mature clinical use—to establish protocols and develop reference tissue phantoms that all will allow universal instrument comparison.
Aim: The largest multi-laboratory comparison of performance assessment in near-infrared diffuse optics is presented, involving 28 instruments and 12 institutions on a total of eight experiments based on three consolidated protocols (BIP, MEDPHOT, and NEUROPT) as implemented on three kits of tissue phantoms. A total of 20 synthetic indicators were extracted from the dataset, some of them defined here anew.
Approach: The exercise stems from the Innovative Training Network BitMap funded by the European Commission and expanded to include other European laboratories. A large variety of diffuse optics instruments were considered, based on different approaches (time domain/frequency domain/continuous wave), at various stages of maturity and designed for different applications (e.g., oximetry, spectroscopy, and imaging).
Results: This study highlights a substantial difference in hardware performances (e.g., nine decades in responsivity, four decades in dark count rate, and one decade in temporal resolution). Agreement in the estimates of homogeneous optical properties was within 12% of the median value for half of the systems, with a temporal stability of <5 % over 1 h, and day-to-day reproducibility of <3 % . Other tests encompassed linearity, crosstalk, uncertainty, and detection of optical inhomogeneities.
Conclusions: This extensive multi-laboratory exercise provides a detailed assessment of near-infrared Diffuse optical instruments and can be used for reference grading. The dataset—available soon in an open data repository—can be evaluated in multiple ways, for instance, to compare different analysis tools or study the impact of hardware implementations.
We present a technique for multispectral fluorescence lifetime imaging with high spatial resolution by combining both single-pixel and data fusion imaging techniques. The system relies on the combined use of three different sensors: two SP cameras capturing multispectral and time-resolved information, and a conventional 2D array detector capturing high spatial resolution images The resultant giga-voxel 4D hypercube is acquired in a fast manner measuring only 0.03% of the dataset. The fusion procedure is done by solving a regularization problem which is efficiently solved via gradient descent. The system can be used to identify fluorophore species.
Significance: Diffuse optical tomography is an ill-posed problem. Combination with ultrasound can improve the results of diffuse optical tomography applied to the diagnosis of breast cancer and allow for classification of lesions.
Aim: To provide a simulation pipeline for the assessment of reconstruction and classification methods for diffuse optical tomography with concurrent ultrasound information.
Approach: A set of breast digital phantoms with benign and malignant lesions was simulated building on the software VICTRE. Acoustic and optical properties were assigned to the phantoms for the generation of B-mode images and optical data. A reconstruction algorithm based on a two-region nonlinear fitting and incorporating the ultrasound information was tested. Machine learning classification methods were applied to the reconstructed values to discriminate lesions into benign and malignant after reconstruction.
Results: The approach allowed us to generate realistic US and optical data and to test a two-region reconstruction method for a large number of realistic simulations. When information is extracted from ultrasound images, at least 75% of lesions are correctly classified. With ideal two-region separation, the accuracy is higher than 80%.
Conclusions: A pipeline for the generation of realistic ultrasound and diffuse optics data was implemented. Machine learning methods applied to a optical reconstruction with a nonlinear optical model and morphological information permit to discriminate malignant lesions from benign ones.
Time-Correlated Single Photon Counting (TCSPC) is a time-resolved and ultrasensitive technique, that provides the analysis of optical pulses to a wide range of different applications both in the biological and chemical domain. Nevertheless, an ultimate constraint to this technique has been historically posed by pile-up distortion, that typically restricts the maximum acquisition speed to few percent of the laser excitation rate. To surpass this fundamental limitation, a novel theoretical solution has been reported in a previous paper: with a perfect matching between detector dead time and laser period, it is possible to achieve a high-speed measurement, still maintaining negligible distortion. In this work, we present the design, characterization and experimental validation of a single-channel TCSPC system that implements the proposed idea. The essential core of the system consists in a compact Detection Head featuring a finely tunable dead time, thanks to a fully-integrated front-end electronics coupled to a custom technology Single-Photon Avalanche Diode (SPAD). This module is providing a picosecond precision timing signal, that is then acquired and digitized by means of a Fast Time to Amplitude Converter (F-TAC) architecture, followed by a high-end Field Programmable Gate Array (FPGA). In order to validate the proposed technique, we carried out on-field fluorescence lifetime measurements employing the newly developed system. The experimental results show good accordance with the previous theoretical framework. It is therefore possible to achieve high acquisition speed (32 Mcps) with an almost null lifetime distortion, thus paving the way to new advanced TCSPC applications.
Multidimensional fluorescence microscopy techniques produce dataset rich of information (space, emission spectrum and lifetime) to investigate photophysical processes in biological samples. To acquire a 4D dataset, one promising microscope design is based on the single-pixel camera scheme and on compressive sensing acquisitions, thanks to which the measurement time can be reduced. Within this framework, a computational step is required to move from the acquisition space to the pixel space and, subsequently, the analysis can be carried out exploiting the high dimensionality. In this work we present an experimental system and a fast-fit method that can produce a map of fluorophore concentrations in parallel to the measurement routine.
A multimodal instrument for breast imaging was developed, combining ultrasound (morphology), shear wave elastography (stiffness), and time domain multiwavelength diffuse optical tomography (blood, water, lipid, collagen) to improve the non-invasive diagnosis of breast cancer.
Multispectral fluorescence lifetime imaging microscopy (λFLIM) is a high sensitivity technique for multifold applications. We present a λFLIM system and a compressive sensing data acquisition strategy, which allows one to reduce the measurement time.
We present a technique to capture high spatial resolution, multispectral, and time-resolved fluorescence images by combining both single-pixel and data fusion imaging techniques. The resultant 4D hypercube can be used to identify fluorophore species.
Performance assessment and standardization are indispensable for instruments of clinical relevance in general and clinical instrumentation based on photon migration/diffuse optics in particular. In this direction, a multi-laboratory exercise was initiated with the aim of assessing and comparing their performances. 29 diffuse optical instruments belonging to 11 partner institutions of a European level Marie Curie Consortium BitMap1 were considered for this exercise. The enrolled instruments covered different approaches (continuous wave, CW; frequency domain, FD; time domain, TD and spatial frequency domain imaging, SFDI) and applications (e.g. mammography, oximetry, functional imaging, tissue spectroscopy). 10 different tests from 3 well-accepted protocols, namely, the MEDPHOT2 , the BIP3 , and the nEUROPt4 protocols were chosen for the exercise and the necessary phantoms kits were circulated across labs and institutions enrolled in the study. A brief outline of the methodology of the exercise is presented here. Mainly, the design of some of the synthetic descriptors, (single numeric values used to summarize the result of a test and facilitate comparison between instruments) for some of the tests will be discussed.. Future actions of the exercise aim at deploying these measurements onto an open data repository and investigating common analysis tools for the whole dataset.
The ability to non-invasively monitor in-vivo the human muscle and adipose tissue is of great practical use and hence of growing interest in the fields of clinical diagnostics and preventive medicine. Optical methods, such as diffuse optical spectroscopy (DOS) applied in the near-infrared spectral region could be of great interest in clinical scenario. In this work, we present a pilot study based on multi-distance broadband time-domain diffuse optical spectroscopy (TD DOS) to characterize in vivo the subcutaneous adipose tissue (abdominal region) and the vastus lateralis muscle (thigh region). The study was performed using a fully automated portable TD DOS instrument on a set of 24 healthy adult volunteers. The optical properties of these two tissue types were obtained over the broad wavelength range of 600-1100 nm. The results suggest a clear influence of the stratified nature of the two regions considered, namely the abdomen and thigh, on the recovered optical properties. This work demonstrates how multi-distance broadband diffuse optical spectroscopy could be complimentary in fields like the non-invasive spectroscopy of adipose tissue and the standard DOS-based muscle oximetry.
To improve non-invasively the specificity in the diagnosis of breast cancer after a positive screening mammography or doubt/suspicious ultrasound examination, the SOLUS project developed a multimodal imaging system that combines: Bmode ultrasound (US) scans (to assess morphology), Color Doppler (to visualize vascularization), shear-wave elastography (to measure stiffness), and time domain multi-wavelength diffuse optical tomography (to estimate tissue composition in terms of oxy- and deoxy-hemoglobin, lipid, water, and collagen concentrations). The multimodal probe arranges 8 innovative photonic modules (optodes) around the US transducer, providing capability for optical tomographic reconstruction. For more accurate estimate of lesion composition, US-assessed morphological priors can be used to guide the optical reconstructions. Each optode comprises: i) 8 picosecond pulsed laser diodes with different wavelengths, covering a wide spectral range (635-1064 nm) for good probing of the different tissue constituents; ii) a large-area (variable, up to 8.6 mm2 ) fast-gated digital Silicon Photomultiplier; iii) the acquisition electronics to record the distribution of time-of-flight of the re-emitted photons. The optode is the basic element of the optical part of the system, but is also a stand-alone, ultra-compact (about 4 cm3 ) device for time domain multi-wavelength diffuse optics, with potential application in various fields.
We have addressed the challenge of investigating the lung using diffuse optics along four different directions, namely: 1) broadband time-domain diffuse optics (TD-DO) in the 600-1100 nm range to derive the mean chest optical properties; 2) Monte Carlo simulations to investigate the depth-sensitivity of TD-DO measurements assuming a layered structure of the chest; 3) single-wavelength TD-DO using a high power pulsed laser on 5 healthy volunteers on a dynamic protocol; 4) single-wavelength TD-DO measurements using a novel large area Silicon Photomultiplier (SIPM) detector module permitting acquisitions at 12 cm source-detector distance.
Multispectral Fluorescence Lifetime Imaging Microscopy (FLIM) is a fundamental tool to study multifold processes in biology and material science. The growing demand for acquisition time reduction requires the parallel acquisition of a multi-dimensional dataset and the exploitation of compressive sensing techniques. In this work we present a multispectral FLIM set-up based on wide-field structured illumination coupled with a spectrometer and a novel time-resolved parallel 18x1 SPAD array detector, working in a single pixel camera scheme. We show the system characterization and its imaging properties varying the compression ratio.
Time-resolved multispectral imaging has recently found many applications ranging from biomedical to environmental field. Multidimensional approach measuring spectral and ultrafast temporal dynamics of fluorescence signal combined with spatial information (imaging) allows one to characterize biological processes at both microscopic and macroscopic level, representing a fundamental step towards development of diagnostic strategies. Long acquisition time is the main drawback of multidimensional approach because it is not compatible with biological system dynamics. In order to reduce the measurement times, it is necessary to parallelize the acquisition (hardware level) and to optimize the acquisition strategy to reduce the measurements number while preserving the information content. In this work we have developed a time-resolved multispectral fluorescence imaging system based on a spad array combined with compression techniques which allows to reduce the number of time-resolved acquisitions by a factor < 70%. The system is based on a double DMD configuration (excitation and detection) coupled to a 32x1 SPAD array, each one with its own TCSPC circuit, placed after an imaging spectrometer. This allows one to use the spatial modulation of the excitation/detection light to acquire images at different wavelengths following the single pixel camera (SPC) scheme. In order to compress the number of acquisitions, a CW fluorescence image is acquired through a CCD and Hadamard transform is applied to select most significative coefficients. The patterns related to these coefficients are subsequently used for SPC acquisition for time and spectral resolution. A Total-Variation based algorithm is used for the reconstruction of the 4D images.
The change in the optical properties of tissue during thermal treatment can be potentially used to monitor procedures like Radiofrequency Ablation (RFA). We present key features in the optical absorption and scattering of tissue during the RFA procedure and during post-ablation cooling down to room temperature. We have used time-resolved diffuse optical spectroscopy for the measurement of the optical properties of tissue for the wavelengths from 650 to 1100 nm. Ex vivo experiments were conducted using a clinical RFA system on bovine liver tissue. Measurements were performed for two temperatures (70°C and 105°C). The following features were observed in the optical properties. First, there was a decrease in optical absorption and an increase in scattering during the treatment. With overtreatment, the absorption increased for initial part of the spectrum (until 910 nm) and scattering decreased in comparison to normal treatment. Secondly, a redshift of the hemoglobin peak and blue shift around water peak was observed in the optical absorption. Finally, a new peak around 840 nm and a valley around 920 nm appeared with heating. When the tissue was allowed to cool down, most of the changes in the absorption around the water peak partially reversed including the blue shift and the valley around 920 nm. Additionally scattering decreased with cooling. Results show key features in the optical properties of tissue during RFA, the effect of overtreatment and post-treatment cooling in ex vivo tissue. Insights from this study will help in advancing optical methods in monitoring thermal treatment.
A widefield system for multidimensional fluorescence imaging capable of resolving space, time and wavelength is developed and validated on a synthetic fluorescence sample. The system enables structured illumination and compressing detection. A compression strategy based on an a-priori information obtained by a camera is validated and proved to be effective at compression ratio of about 90%.
In the last decade, multimodal imaging raised increasing interest to overcome the limits of single techniques and improve the diagnostic potential during the same examination. This gives rise to the need for phantoms and procedures for standardizing performance assessment of the multimodal instrument. The SOLUS1 project adopts this methodology with the aim to build a multimodal instrument (based on diffuse optics -DO-, shear wave elastography -SWE-, and ultrasound imaging -US-) to increase the specificity of breast cancer diagnosis. Here we propose a long-lasting phantom based on silicone material (easier to manipulate with respect to other material for bimodal phantom such as polyvinyl alcohol, PVA) and suitable for both diffuse optical imaging/tomography and ultrasound acquisitions, designed within the SOLUS project. To achieve this goal, we explored a new silicone material for diffuse optics and ultrasound (Ecoflex 00-30), creating a new fabrication recipe and demonstrating its suitability for multimodal imaging if coupled to another silicone elastomer (Sylgard 184), featuring similar optical and acoustical performances except for the echogenicity. The main advantage of the proposed phantom is the capability of tuning independently optical and acoustical performances, thus allowing one to mimic a wide range of clinical scenarios.
Diffuse Optical Tomography (DOT) is a tool for 3D reconstruction of absorption and scattering inside a tissue. Typically, this method requires a dense distribution of sources and detectors, thus hampering the possibility of fully exploring a time-resolved detection. Recently, techniques based on structured-light illumination and compressing detection have been developed, opening the possibility of fully exploiting a source/detector spatial modulation for compression at the measurement stage. Here we propose a combined Continuous-Wave (CW) and time-domain (TD) adaptive scheme based on the singular-value decomposition (SVD) for optimal-patterns calculation. Patterns are firstly computed based on a fast acquisition via a CCD, and consequently projected for time-resolved measurements.
Time Domain Diffuse Optical Tomography (TD-DOT) performed at multiple wavelengths can be used to non-invasively probe tissue composition. Then, tissue composition can be related to breast tissue and lesion type. Thus, TD-DOT could be used for therapy monitoring for breast cancer. We developed a software tool for multi-wavelength TD-DOT and performed a validation on meat phantoms to mimic tissue heterogeneity. An inclusion of different meat was exploited to mimic the presence of a lesion in the breast. Results show good localization of the inclusion, but poor quantification of the reconstructed breast composition. The use of a morphological prior constraint, providing information on inclusion geometry and position, significantly improves both localization and composition estimate.
Diffuse optical imaging can be used to probe highly scattering media like biological tissue down to a depth of few centimeters, with spatial resolution limited by light scattering. Its combination with ultrasound imaging can potentially lead to medical imaging systems with, for instance, high specificity in the examination of tumors. However, the presence of the ultrasound coupling gel between probe and tissue can have detrimental effects on the accuracy of optical imaging techniques. Here we present an experimental study on the effect of ultrasound coupling fluids on diffuse optical spectroscopy (DOS) and diffuse correlation spectroscopy (DCS). We demonstrate on tissue-mimicking phantoms that the use of standard water-clear gels, providing a direct path for the light from the source to the detection point, can distort optical measurements generating strong underestimation of both the absorption and the reduced scattering coefficients in DOS measurements, as well as underestimation of the Brownian diffusion coefficient in DCS measurements. On the contrary, various turbid fluids demonstrate excellent performance in preventing this issue.
Radiofrequency ablation (RFA) is minimally invasive thermotherapy, where a heating source is used to target and kill malignant cells in a tissue. While RFA has tremendous potential in the field of oncology, there is also a need for reliable real-time monitoring of this procedure to avoid over or under treatment. In this work, we investigate the use of timeresolved diffuse optical spectroscopy (DOS) to continuously track the change in optical properties during RFA to monitor the process of ablation. The time evolution of the spectra of the optical properties of the tissue undergoing treatment gives deep insights into the structural and constitutional changes occurring during the RFA treatment.
For the first time, we are proposing a compressive-sensing approach to time-domain diffuse Raman spectroscopy for depth probing of multilayer diffusive media. We built a spectrometer capable of both spectral and temporal acquisition with a single-pixel detector and tested it for depth sectioning on a bilayer tissue-mimicking phantom.
Time Domain Diffuse Optical Tomography (TD-DOT) at different wavelengths can be used to retrieve tissue reconstructing the components of a two-region system starting from self-normalized time-dependent measurements performed in reflectivity geometry over multiple wavelengths. The proposed method performs a fit of a limited number of tissues parameters providing a good quantification of the components’ concentrations by applying a FEM-based Diffusion approximation of the TD-DOT direct model.
Time-resolved imaging is a valuable tool for biomedical applications such as Diffused Optical Tomography (DOT) and Fluorescence Lifetime Imaging (FLIM). The first one characterizes and localizes absorption/scattering heterogeneities, which can be representative of tumors, and is routinely used for brain functional imaging. FLIM provides relevant information (e.g. pH, ion concentration and FRET) in cell biology and find application in molecular imaging for preclinical studies in small animals. Beyond biomedical applications, time-resolved imaging is exploited for environmental monitoring, LIDAR and characterization of combustion processes.
Structured light illumination and compressive-sensing detection have been recently proposed as new strategies to preserve information content while significantly reducing the number of measurements. One possible implementation of this approach is the Single Pixel Camera (SPC), where the inner product between the image of the subject and appropriate patterns is measured by using a spatial modulator (e.g. DMD, SLM) and focusing the light on a single pixel detector.
In this work, we present a time-resolved imaging system for DOT applications based on structured light illumination and SPC detection, implementing an adaptive scheme based on Singular-Value Decomposition for optimal generation of input/output patterns. Moreover, a novel scheme of time-resolved camera, with ps temporal resolution, is proposed and experimentally validated. The device consists of a high-density matrix of single photon detection elements which can be selectively enabled/disabled. Spatial modulator and detector are combined into a single chip improving cost and compactness. In conclusion, the proposed time-resolved imaging approach can have significant impact on biomedical, environmental and LIDAR applications as an alternative to gated cameras or scanning systems.
Time-resolved imaging is a fundamental tool for biomedical applications such as fluorescence lifetime imaging microscopy (FLIM) and mapping of tissue optical parameters. FLIM, in particular, enables us to study the micro-environment of fluorophores in cell biology, providing relevant information like pH, ion concentration and molecular coupling (e.g. FRET).
Compressed sensing approaches, based on image sparsity, have been recently proposed as a novel imaging paradigm allowing to preserve information content while significantly reducing the number of measurements. Single Pixel Camera (SPC) approach is one possible implementation of this idea. The object is imaged on a spatially modulated system (e.g. DMD, SLM), then by focusing the exiting light on a single pixel detector, the inner product between image and pattern is measured.
In this work we propose and validate a novel scheme of time-resolved camera with ps temporal resolution where all the elements required for compressed sensing are combined into a single chip, allowing a significant cost reduction, compactness and performance improvement. The proposed device is based on a high-density array of detection elements, operating in the single photon regime, which can be selectively enabled/disabled. All pixels are connected to one single Time-to-Digital Converter (TDC). In order to experimentally validate the imaging and temporal capability of the proposed system, fluorescence lifetime imaging and time-gated imaging in a diffused medium have been carried out. We believe the proposed time-resolved camera can be a convenient approach in many biomedical applications where a gated camera or a time-resolved scanning system are currently used.
Time domain Diffuse Optical Tomography (TD-DOT) non-invasively probes the optical proprieties of biological tissue. These can be related to changes in tissue composition, thus making TD-DOT potentially valuable for cancer imaging. In particular, an application of interest is therapy monitoring for breast cancer. Thus, we developed a software tool for multiwavelength TD-DOT in reflectance geometry. While the use of multiple wavelengths probes the main components of the breast, the chosen geometry offers the advantage of linking the photon flight time to the investigated depth. We validated the tool on silicon phantoms embedding an absorbing inclusion to simulate a malignant lesion in breast tissue. Also, we exploited the a priori information on position and geometry of the inclusion by using a morphological prior constraint. The results show a good localization of the depth of inclusion but a reduced quantification. When the morphological constraint is used, though, the localization improves dramatically, also reducing surface artifacts and improving quantification as well. Still, there is room for improvement in the quantification of the “lesion” properties.
The use of bioresorbable fibers represents an innovative way to build optical implantable devices and to look inside the body. Recently, a new kind of bioresorbable fibers, based on calcium-phosphate glasses, has been introduced by some of us. They show a good biocompatibility and improved attenuation loss coefficient with respect to other bioresorbable fibers. In this work, we used those fibers to explore their suitability in diffuse optics. Indeed, the time-domain technique is a non-invasive methodology which allows to have an absolute estimate of the absorption and reduced scattering spectra of the diffusive medium. It allows to bring information about concentration of chemical components (water, oxyand deoxy-hemoglobin), thus conveying information about the functional status and/or the scattering properties (changes in tissue microstructure, edema). Such information can then be related to the tissue regeneration, healing process, or to a harmful evolution. This makes the time domain optical spectroscopy coupled to bioresorbable fibers a good candidate for future medical devices. Here we demonstrate the suitability of these fibers for diffuse optics by means of standardized tests and then we use them for a proof-of-principle measurement on ex-vivo chicken breast, obtaining results comparable with standard fibers. Thanks to the encouraging results, we are working on a system based on a single fiber (serving as both injection and collection fiber) to go closer to a single interstitial fiber which can lessen the effect of the implant.
Diffuse Optical Tomography (DOT)is a powerful tool for the reconstruction of optical properties inside a diffusive medium, such as biological tissues. In particular, in the last years, techniques based on structured light illumination and compressive sensing detection have been developed. In this work a time-resolved system based on structured light illumination and compressive detection has been developed and used for DOT. Moreover, a data-driven algorithm for optimal pattern generation based on the Singular-Value Decomposition has been implemented and validated.
KEYWORDS: Absorption, Scattering, Tissues, Diffuse optical spectroscopy, Data modeling, Tissue optics, In vivo imaging, Optical properties, Abdomen, Medicine
A periodic monitoring of the adipose tissue functions due to interventions, such as calorie restriction and bariatric surgery, or pathophysiological processes, has an increasing relevance in clinical diagnostics. Diffuse Optical Spectroscopy (DOS) is a valuable non-invasive tool that can be used in that direction. In this work, we present a pilot study based on Time Domain Broadband Diffuse Optical Spectroscopy (TD DOS) to characterize in vivo the subcutaneous fat tissue in the abdominal region. A first of its kind, portable TD DOS instrumentation, already enrolled in clinical studies, was used. Three healthy male volunteers were considered. Three source-detector separation distances (1, 2, and 3 cm) were used over the broad wavelength range of 600-1100 nm. The analysis was performed using a method based on a heterogeneous model to account for the multi-layered nature of the subcutaneous adipose tissue, and to obtain the optical properties specific to this fat localization. Inter-subject variation of tissue composition data was observed.
Non-contact scanning at small source-detector separation enables imaging of cerebral and extracranial signals at high spatial resolution and their separation based on early and late photons accounting for the related spatio-temporal characteristics.
A time-resolved Diffuse Optical Tomography system based on multiple view
acquisition, pulsed structured light illumination and detection with spatial compression is
proposed. Reconstructions on heterogeneous tissue mimicking phantoms are presented.
In this work an analytical model for the time-resolved signal emitted by a
uniformly distributed Raman scatterer in a diffusive parallelepiped is derived and validated
with Monte Carlo (MC) simulations.
To evaluated capabilities of multispectral TD-DOT systems in reflection geometry, we
performed a measurement campaign on multimaterial composition phantoms. Results show correct
composition gradation of inclusions but still lack absolute accuracy.
We present the first broadband (600-1100 nm) diffuse optical characterization of thyroglobulin and tyrosine, which are thyroid-specific tissue constituents. In-vivo measurements at the thyroid region enabled their quantification for functional and diagnostic applications.
We show for the first time the aptness of Calcium Phosphate Glass-based bioresorbable fibers for time-domain diffuse optics using tests described by a standardized protocol and we also present a spectroscopic measurement on a chicken breast.
We investigated depth heterogeneity in the abdomen using time-domain diffuse optical spectroscopy at 3 source-detector distances, finding a higher water content in shallower regions, possibly ascribed to fat heterogeneity and/or skin contributions.
Diffuse Optical Tomography (DOT) can be described as a highly multidimensional problem generating a huge data set with long acquisition/computational times. Biological tissue behaves as a low pass filter in the spatial frequency domain, hence compressive sensing approaches, based on both patterned illumination and detection, are useful to reduce the data set while preserving the information content. In this work, a multiple-view time-domain compressed sensing DOT system is presented and experimentally validated on non-planar tissue-mimicking phantoms containing absorbing inclusions.
We propose to couple a single-pixel camera with a photon counting board in order to obtain an inexpensive time-resolved imaging system having a high spatial and temporal resolution. As an alternative to compressive sensing, a wavelet-based adaptive acquisition strategy is employed which allows high compression rates with little degradation of the image quality. The applicability of our approach is demonstrated for fluorescence lifetime imaging. Experimental results obtained by imaging samples embedding several fluorophores are provided. The proposed imaging system with the wavelet-based strategy can be suitable for a microscope in order to perform fluorescence lifetime imaging microscopy measurements.
The study of photon migration through highly scattering media opens the way to the non-invasive investigation of biological tissues well below the skin surface. When the medium is addressed in reflectance geometry, a key issue is to maximize the depth reached by migrating photons. By exploiting the Diffusion Approximation of the Radiative Transfer Equation, we calculated the time-resolved and continuous-wave probability density functions for the maximum depth reached by detected photons, for both a homogeneous and a layered laterally-infinite diffusive slab. From the probability density functions it is possible to calculate the mean value of the maximum depth at which detected photons have undergone scattering events.
Reduction in scattering, high absorption, and spectral features of tissue constituents above 1000 nm could help in gaining higher spatial resolution, penetration depth, and specificity for in vivo studies, opening possibilities of near-infrared diffuse optics in tissue diagnosis. We present the characterization of collagen absorption over a broadband range (500 to 1700 nm) and compare it with spectra presented in the literature. Measurements were performed using a time-domain diffuse optical technique. The spectrum was extracted by carefully accounting for various spectral distortion effects, due to sample and system properties. The contribution of several tissue constituents (water, lipid, collagen, oxy, and deoxy-hemoglobin) to the absorption properties of a collagen-rich in vivo bone location, such as radius distal in the 500- to 1700-nm wavelength region, is also discussed, suggesting bone diagnostics as a potential area of interest.
The recent developments in time-domain diffuse optics that rely on physical concepts (e.g., time-gating and null distance) and advanced photonic components (e.g., vertical cavity source-emitting laser as light sources, single photon avalanche diode, and silicon photomultipliers as detectors, fast-gating circuits, and time-to-digital converters for acquisition) are focused. This study shows how these tools could lead on one hand to compact and wearable time-domain devices for point-of-care diagnostics down to the consumer level and on the other hand to powerful systems with exceptional depth penetration and sensitivity.
A mechanically switchable solid inhomogeneous phantom simulating localized absorption changes was developed and characterized. The homogeneous host phantom was made of epoxy resin with black toner and titanium dioxide particles added as absorbing and scattering components, respectively. A cylindrical rod, movable along a hole in the block and made of the same material, has a black polyvinyl chloride cylinder embedded in its center. By varying the volume and position of the black inclusion, absorption perturbations can be generated over a large range of magnitudes. The phantom has been characterized by various time-domain diffuse optics instruments in terms of absorption and scattering spectra, transmittance images, and reflectance contrast. Addressing a major application of the phantom for performance characterization for functional near-infrared spectroscopy of the brain, the contrast was measured in reflectance mode while black cylinders of volumes from ≈20 mm3 to ≈270 mm3 were moved in lateral and depth directions, respectively. The new type of solid inhomogeneous phantom is expected to become a useful tool for routine quality check of clinical instruments or implementation of industrial standards provided an experimental characterization of the phantom is performed in advance.
We report on the design, performance assessment, and first in vivo measurement of a Time-Resolved Diffuse Optical system for broadband (600-1350 nm) nm measurement of absorption and scattering spectra of biological tissues for non-invasive clinical diagnostics. Two strategies to reduce drift and enhance responsivity are adopted. The system was enrolled in a first in vivo test phase on healthy volunteers, carrying out non-invasive, in vivo quantification of key tissue constituents (oxy- and deoxy-hemoglobin, water, lipids, collagen) and tissue micro-structure (scatterer size and density).
We report a broadband time-resolved characterization of selected bony prominence locations of the human body. A clinical study was performed at six different bony prominence locations of 53 subjects. A portable broadband time-resolved system equipped with pulse drift and distortion compensation strategy was used for absorption and scattering measurements. Key tissue constituents were quantified as a pilot step towards non-invasive optical assessment of bone pathologies.
We propose a simple and reliable solid phantom for mimicking realistic localized absorption changes within a diffusive medium. The phantom is based on a solid matrix holding a movable black inclusion embedded in a rod. Translating the rod parallel to the phantom surface, the inhomogeneity can be positioned beneath the source-detector pair (perturbed case) or far from it (unperturbed case). Examples of time-resolved transmittance measurements and time-resolved reflectance scans are shown to demonstrate the properties and the versatility of the phantom.
Diffuse optics is a powerful tool for clinical applications ranging from oncology to neurology, but also for molecular imaging, and quality assessment of food, wood and pharmaceuticals. We show that ideally time-domain diffuse optics can give higher contrast and a higher penetration depth with respect to standard technology. In order to completely exploit the advantages of a time-domain system a distribution of sources and detectors with fast gating capabilities covering all the sample surface is needed. Here, we present the building block to build up such system. This basic component is made of a miniaturised source-detector pair embedded into the probe based on pulsed Vertical-Cavity Surface-Emitting Lasers (VCSEL) as sources and Single-Photon Avalanche Diodes (SPAD) or Silicon Photomultipliers (SiPM) as detectors. The possibility to miniaturized and dramatically increase the number of source detectors pairs open the way to an advancement of diffuse optics in terms of improvement of performances and exploration of new applications. Furthermore, availability of compact devices with reduction in size and cost can boost the application of this technique.
Time-resolved diffuse optical spectroscopy provides non-invasively the optical characterization of highly diffusive media, such as biological tissues. Light pulses are injected into the tissue and the effects of light propagation on re-emitted pulses are interpreted with the diffusion theory to assess simultaneously tissue absorption and reduced scattering coefficients. Performing spectral measurements, information on tissue composition and structure is derived applying the Beer law to the measured absorption and an empiric approximation to Mie theory to the reduced scattering. The absorption properties of collagen powder were preliminarily measured in the range of 600-1100 nm using a laboratory set-up for broadband time-resolved diffuse optical spectroscopy. Optical projection images were subsequently acquired in compressed breast geometry on 218 subjects, either healthy or bearing breast lesions, using a portable instrument for optical mammography that operates at 7 wavelengths selected in the range 635-1060 nm. For all subjects, tissue composition was estimated in terms of oxy- and deoxy-hemoglobin, water, lipids, and collagen. Information on tissue microscopic structure was also derived. Good correlation was obtained between mammographic breast density (a strong risk factor for breast cancer) and an optical index based on collagen content and scattering power (that accounts mostly for tissue collagen). Logistic regression applied to all optically derived parameters showed that subjects at high risk for developing breast cancer for their high breast density can effectively be identified based on collagen content and scattering parameters. Tissue composition assessed in breast lesions with a perturbative approach indicated that collagen and hemoglobin content are significantly higher in malignant lesions than in benign ones.
Diffuse Optical Tomography (DOT) and Fluorescence Molecular Tomography (FMT) generally require a huge data set which poses severe limits to acquisition and computational time, especially with a multidimensional data set. The highly scattering behavior of biological tissue leads to a low bandwidth of the information spatial distribution and hence the sampling can be preferably carried out in the spatial frequency source/detector space. In this work, a time-resolved single pixel camera scheme combined with structured light illumination is presented and experimentally validated on phantoms measurements. This approach leads to a significant reduction of the data set while preserving the information content.
We propose a simple and reliable solid phantom for mimicking localized absorption changes within a diffusive medium. The phantom is based on the Equivalence Relation stating that any realistic absorption inhomogeneity can be mimicked by a totally absorbing sphere of adequate volume. Applying this concept, we constructed a solid phantom holding a movable black inclusion to be positioned beneath the source-detector pair (perturbed case) or far from it (unperturbed case). Different absorption perturbations can be mimicked by changing the volume and the position of the black object both in transmittance and reflectance configuration. Time-resolved measurements of transmittance images and a lateral reflectance scan are presented.
Diffuse Optics is growing in terms of applications ranging from e.g. oximetry, to mammography, molecular imaging, quality assessment of food and pharmaceuticals, wood optics, physics of random media. Time-domain (TD) approaches, although appealing in terms of quantitation and depth sensibility, are presently limited to large fiber-based systems, with limited number of source-detector pairs. We present a miniaturized TD source-detector probe embedding integrated laser sources and single-photon detectors. Some electronics are still external (e.g. power supply, pulse generators, timing electronics), yet full integration on-board using already proven technologies is feasible. The novel devices were successfully validated on heterogeneous phantoms showing performances comparable to large state-of-the-art TD rack-based systems. With an investigation based on simulations we provide numerical evidence that the possibility to stack many TD compact source-detector pairs in a dense, null source-detector distance arrangement could yield on the brain cortex about 1 decade higher contrast as compared to a continuous wave (CW) approach. Further, a 3-fold increase in the maximum depth (down to 6 cm) is estimated, opening accessibility to new organs such as the lung or the heart. Finally, these new technologies show the way towards compact and wearable TD probes with orders of magnitude reduction in size and cost, for a widespread use of TD devices in real life.
Performance assessment of instruments devised for clinical applications is of key importance for validation and quality assurance. Two new protocols were developed and applied to facilitate the design and optimization of instruments for time-domain optical brain imaging within the European project nEUROPt. Here, we present the “Basic Instrumental Performance” protocol for direct measurement of relevant characteristics. Two tests are discussed in detail. First, the responsivity of the detection system is a measure of the overall efficiency to detect light emerging from tissue. For the related test, dedicated solid slab phantoms were developed and quantitatively spectrally characterized to provide sources of known radiance with nearly Lambertian angular characteristics. The responsivity of four time-domain optical brain imagers was found to be of the order of 0.1 m2 sr. The relevance of the responsivity measure is demonstrated by simulations of diffuse reflectance as a function of source-detector separation and optical properties. Second, the temporal instrument response function (IRF) is a critically important factor in determining the performance of time-domain systems. Measurements of the IRF for various instruments were combined with simulations to illustrate the impact of the width and shape of the IRF on contrast for a deep absorption change mimicking brain activation.
The optical properties of the human head in the range from 600 nm to 1100 nm have been non-invasively in-vivo investigated by various research groups using different diffuse optics techniques and data analysis methods.
Three recipes for tissue constituent-equivalent phantoms of water and lipids are presented. Nature phantoms are made using no emulsifying agent, but just a professional disperser, instead Agar and Triton phantoms are made using agar or Triton X-100, respectively, as agents to emulsify water and lipids. Different water-to-lipid ratios ranging from 30 to 70 percent by mass are proposed and tested. Optical characterization by time-resolved spectroscopy was performed in terms of optical properties, homogeneity, reproducibility and composition retrieval.
KEYWORDS: Lung, Absorption, Monte Carlo methods, Tissue optics, Chest, In vivo imaging, Optical properties, Scattering, Optical spectroscopy, Reflectivity
Monte Carlo simulations and preliminary time-resolved spectroscopy measurements were performed to investigate the feasibility of the in vivo optical diagnostics of lung conditions and diseases. Absorption and reduced scattering properties of the chest, arising from in vivo spectral measurements on volunteers are presented.
Presently time-resolved optical spectroscopy is applied with increasing success for non-invasive medical diagnostics mainly up to 1100 nm. We extended the investigation range beyond this limit, employing a supercontinuum fiber laser source and a Single-Photon Avalanche Diode in InGaAs/InP operated in gated mode. First in-vivo measurements were performed on the forearm and the breast of two healthy volunteers, reaching up to 1360 nm.
Time-domain diffuse optical spectroscopy has become a powerful tool to study highly scattering media, mainly in the
fields of non-invasive medical diagnostics and quality assessment of food and pharmaceutical products. Up to now this
technique has been exploited mostly up to 1100 nm: we extend the spectral range by means of a continuously tunable
pulsed laser source at a high repetition rate and a custom InGaAs/InP Single-Photon Avalanche Diode operated in time-gated
mode, working up to 1700 nm. The characterization of the system is presented. As a first example of application,
we measured the absorption spectrum of collagen powder in the range 1100 - 1700 nm, which could prove useful for
breast density assessment.
We propose a system for time-domain diffuse optical spectroscopy extending up to 1300 nm based on a supercontinuum
source and time-correlated single photon counting detection. The system was validated on liquid
phantoms demonstrating a good linearity up to an absorption coefficient of 2.0 cm-1. Preliminary measurements
on collagen powder reveal an important absorption peak around 1200 nm. Feasibility of in vivo diffuse optical
spectroscopy on the female breast is demonstrated up to 1200 nm at an interfiber distance of 2 cm.
Spectral measurements were performed from 900 to 1300 nm, using a fully automated set-up for time domain optical
spectroscopy. Spectrally selected picosecond pulses emitted from a supercontinuum fiber source were used for
illumination. The detection of re-emitted pulses was achieved using a photomultiplier tube with InP/InGaAsP
photocathode, followed by a PC board for time-correlated single photon counting.
To allow the estimate of tissue composition at long wavelengths, the optical characterization of collagen type I powder
was extended up to 1300 nm. A marked absorption peak was detected around 1200 nm, which could prove useful for
collagen quantification from in vivo optical data.
In vivo spectral measurements of breast tissue were performed for the first time from 900 to 1300 nm in reflectance
geometry. The sensitivity of the detector was very low above 1200 nm, still it allowed us to reveal a long-wavelength
range (1000-1300 nm) potentially interesting for applications. A dominant absorption peak is present around 1200 nm.
All major tissue constituents (i.e., water, lipid, and collagen) contribute to it. Thus, it is potentially interesting for the
assessment of tissue composition, but it might cause exceeding attenuation in some practical cases. However, slightly
shorter wavelengths (i.e. 1100-1150 nm) corresponding to the raising edge of the peak, might allow an accurate estimate
of tissue composition, with the advantage of much lower attenuation.
Cardiovascular diseases are the main cause of death in industrialized countries. Worldwide, a large number of
patients suffering from cardiac diseases are treated by surgery. Despite the advances achieved in the last decades
with myocardial protection, surgical failure can still occur. This is due at least in part to the imperfect control
of the metabolic status of the heart in the various phases of surgical intervention. At present, this is indirectly
controlled by the electrocardiogram and the echographic monitoring of cardiac mechanics as direct measurements
are lacking. Diffuse optical technologies have recently emerged as promising tools for the characterization of
biological tissues like breast, muscles and bone, and for the monitoring of important metabolic parameters such
as blood oxygenation, volume and flow. As a matter of fact, their utility has been demonstrated in a variety of
applications for functional imaging of the brain, optical mammography and monitoring of muscle metabolism.
However, due to technological and practical difficulties, their potential for cardiac monitoring has not yet been
exploited. In this work we show the feasibility of the in-vivo determination of absorption and scattering spectra
of the cardiac muscle in the 600-1100 nm range, and of monitoring myocardial tissue hemodynamics by time
domain near-infrared spectroscopy at 690 nm and 830 nm. Both measurements have been performed on the
exposed beating heart during open chest surgery in pigs, an experimental model closely mimicking the clinical
cardio-surgical setting.
Time domain diffuse optical spectroscopy provides the absorption and scattering properties of biological tissues
and diffusive materials. Few measurements are available at discrete wavelengths beyond 1100 nm, and just one
time-domain system continuously tuneable up to 1400 nm. We developed a time-domain system, based on a
continuously tuneable supercontinuum pulsed source, and a custom InGaAs/InP Single-Photon Avalanche Diode.
Operation was demonstrated in the 1100-1700 nm range with a spectral resolution of 15 nm, a temporal resolution
of 150 ps and a background of 6000 counts/s. A first example of application on the optical characterization of
collagen powder is given.
To facilitate the design and optimization of instruments for time-domain optical brain imaging within the European
project "nEUROPt", the performance of various instruments is assessed and compared. This type of instruments relies on
picosecond lasers with high repetition rates, fast detectors and time-correlated single photon counting. The first step of
the assessment included a number of basic tests that are related to parameters of the source, to the differential
nonlinearity of the timing electronics and to the temporal instrument response function (IRF). An additional test has been
devised to measure the responsivity of the detection system, i.e. the overall efficiency to collect and detect light
emerging from tissue. Dedicated solid slab phantoms have been developed and quantitatively spectrally characterized to
provide sources of known radiance with nearly Lambertian angular characteristics. The wavelength-dependent
transmittance factor of these phantoms was of the order of 1020/(W s m2sr). Measurements of the responsivity of the
detection systems of three time-domain optical brain imagers tested yielded similar values of the order of 0.1 mm2sr.
The feasibility of in vivo measurements in the range of 1000 to 1100 nm and the potential benefits of operation in that wavelength range for diagnostic applications are investigated. To this purpose, an existing system for time-resolved diffuse spectroscopy is modified to enable in vivo studies to be carried out continuously from 600 to 1100 nm. The optical characterization of collagen powder is extended to 1100 nm and an accurate measurement of the absorption properties of lipid is carried out over the entire spectral range. Finally, the first in vivo absorption and scattering spectra of breast tissue are measured from 10 healthy volunteers between 600 and 1100 nm and tissue composition is evaluated in terms of blood parameters and water, lipid, and collagen content using a spectrally constrained global fitting procedure.
We discuss the spectral distortions occurring when time-resolved diffuse spectroscopy is performed illuminating
with a spectrally wide source. We show that the spectral region within the source bandwidth that exhibits
the lowest absorption will dominate the resulting time-resolved curve, leading to significant distortions on the
retrieved absorption spectrum (including shifts in peak positions). Due to the nonlinear behavior of the light
attenuation due to absorption, this effect becomes more pronounced when including longer and longer photon
path lengths. First, a theoretical treatment of the problem is given and then the distortion is described by timeresolved
reflectance simulations and experimental measurements of lipid and water samples. Finally, a spectrally
constrained data analysis is proposed to overcome the distortion and improve the accuracy of the estimation of
chromophore concentrations from absorption spectra. Measurements on a lipid sample show a reduction of the
error from 30% to 6%.
KEYWORDS: Scattering, Absorption, Monte Carlo methods, Optical properties, Data modeling, Picosecond phenomena, Signal to noise ratio, Near infrared, Photon counting, Geometrical optics
In this work we studied the accuracy of a non-linear fitting procedure, based on the Levenberg-Marquardt algorithm,
for time-resolved measurements to retrieve the absorption and the reduced scattering coefficients of
an absorbing diffusive medium. This procedure is suitable for retrieving optical properties in a wider range of
situations (e.g. solid samples, reflectance geometry), with respect to the linear inversion procedures recently
presented for both CW and time domain measurements. By means of both analytical and numerical (Monte
Carlo) simulations, we quantified the influence of photon counts, temporal sampling, analytical model, background
and instrument response function on the accuracy in the estimation of the optical properties. Also a new
analytical model to describe light propagation in diffusive media based on the Radiative Transport Equation has
been considered. The main source of error that affects the accuracy of the absorption and reduced scattering
coefficients retrieved by the non-linear procedure appears to be the analytical model adopted in the inversion
procedure.
We describe the development of a compact time-resolved system for the measurement of the optical properties of highly scattering media over a bandwidth of 600-1100 nm. The instrument is based on a fiber laser generating supercontinuum radiation, that is spectrally dispersed and used to sequentially illuminate the sample. A single photon avalanche photo-diode in combination with time correlated single-photon counting is used to recover the time-dispersion curve at each wavelength. The calibration of the system and in-vivo applications are shown.
Time-resolved transmittance measurements were performed in the wavelength range of 610 or 700 to 1050 nm on
phantom slabs and bone tissue cubes of different sizes. The data were best fitted with solutions of the diffusion equation
for an infinite slab and for a parallelepiped to investigate how size and optical properties of the samples affect the results
obtained with the two models. When small samples are considered, the slab model overestimates both optical
coefficients, especially the absorption. The parallelepiped model largely compensates for the small sample size and
performs much better also when the absorption spectra are interpreted with the Beer's law to estimate bone tissue
composition.
In this work we propose and demonstrate that time-resolved optical spectroscopy in the spectral region 700-1040 nm, on
a picosecond time scale, is a valuable technique for non-invasive wood characterization. Two different wood types have
been considered, fir and oak chestnut as an example of softwood and hardwood, respectively. Wood samples have been
measured in three different conditions: dry, wet and degraded by an ozone treatment. The two types of wood show
different absorption and scattering spectra according to the treatment, revealing both chemical and structural changes.
In spite of many progresses achieved both with theories and with experiments in studying light propagation through
diffusive media, a reliable method for accurate measurements of the optical properties of diffusive media at NIR
wavelengths is, in our opinion, still missing. It is therefore difficult to create a reference diffusive medium. We describe
two methods in the CW and time domain to calibrate the reduced scattering coefficient of a liquid diffusive medium and
the absorption coefficient of an absorber with a standard error smaller than 2% for both the coefficients.
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