Initial non-invasive in vivo sensing of the lung using time domain diffuse optics.

The in vivo diagnosis and monitoring of pulmonary disorders (caused for example by emphysema, Covid-19, immature lung tissue in infants) could be effectively supported by the non-invasive sensing of the lung through light. With this purpose, we investigated the feasibility of probing the lung by means of time-resolved diffuse optics, leveraging the increased depth (a few centimeters) attained by photons collected after prolonged propagation time (a few nanoseconds). We present an initial study that includes measurements performed on 5 healthy volunteers during a breathing protocol, using a time-resolved broadband diffuse optical spectroscopy system. Those measurements were carried out across the spectral range of 600-1100 nm at a source-detector distance of 3 cm, and at 820 nm over a longer distance (7-9 cm). The preliminary analysis of the in vivo data with a simplified homogeneous model revealed a maximum probing depth of 2.6-3.9 cm, suitable for reaching the lung. Furthermore, we observed variations in signal associated with respiration, particularly evident at long photon propagation times. However, challenges stemming from both intra- and inter-subject variability, along with inconsistencies potentially arising from conflicting scattering and absorption effects on the collected signal, hindered a clear interpretation. Aspects that require further investigation for a more comprehensive understanding are outlined.

Time domain diffuse Raman spectroscopy using single pixel detection.

Diffuse Raman spectroscopy (DIRS) extends the high chemical specificity of Raman scattering to in-depth investigation of thick biological tissues. We present here a novel approach for time-domain diffuse Raman spectroscopy (TD-DIRS) based on a single-pixel detector and a digital micromirror device (DMD) within an imaging spectrometer for wavelength encoding. This overcomes the intrinsic complexity and high cost of detection arrays with ps-resolving time capability. Unlike spatially offset Raman spectroscopy (SORS) or frequency offset Raman spectroscopy (FORS), TD-DIRS exploits the time-of-flight distribution of photons to probe the depth of the Raman signal at a single wavelength with a single source-detector separation. We validated the system using a bilayer tissue-bone mimicking phantom composed of a 1 cm thick slab of silicone overlaying a calcium carbonate specimen and demonstrated a high differentiation of the two Raman signals. We reconstructed the Raman spectra of the two layers, offering the potential for improved and quantitative material analysis. Using a bilayer phantom made of porcine muscle and calcium carbonate, we proved that our system can retrieve Raman peaks even in the presence of autofluorescence typical of biomedical tissues. Overall, our novel TD-DIRS setup proposes a cost-effective and high-performance approach for in-depth Raman spectroscopy in diffusive media.

Fast time-domain diffuse correlation spectroscopy with superconducting nanowire single-photon detector: system validation and in vivo results.

Time-domain diffuse correlation spectroscopy (TD-DCS) has been introduced as an advancement of the "classical" continuous wave DCS (CW-DCS) allowing one to not only to measure depth-resolved blood flow index (BFI) but also to extract optical properties of the measured medium without using any additional diffuse optics technique. However, this method is a photon-starved technique, specially when considering only the late photons that are of primary interest which has limited its in vivo application. In this work, we present a TD-DCS system based on a superconducting nanowire single-photon detector (SNSPD) with a high quantum efficiency, a narrow timing response, and a negligibly low dark count noise. We compared it to the typically used single-photon avalanche diode (SPAD) detector. In addition, this system allowed us to conduct fast in vivo measurements and obtain gated pulsatile BFI on the adult human forehead.

Interstitial null-distance time-domain diffuse optical spectroscopy using a superconducting nanowire detector.

Significance: Interstitial 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. Aim: To 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). Approach: In particular, we demonstrate the use of a superconducting nanowire single photon detector (SNSPD) to perform TD-DOS at almost NSDS ( ≈ 150 µ m ) 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. Results: This 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. Conclusions: Utilizing 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.

Solid optical tissue phantom tools based on upconverting nanoparticles for biomedical applications.

Significance: Phantoms play a critical role in the development of biophotonics techniques. There is a lack of novel phantom tools in the emerging field of upconverting nanoparticles (UCNPs) for biophotonics application. This work provides a range of UCNP-based phantom tools and a manufacturing recipe to bridge the gap and accelerate the development of UCNP-based biophotonics applications. Aim: The study aims to provide a well-characterized UCNP-based solid phantom recipe and set of phantom tools to address a wide range of UCNP-based biophotonics applications. Approach: A solid phantom recipe based on silicone matrix was developed to manufacture UCNP-based phantoms. A lab built UCNP imaging system was used to characterize upconverted fluorescence emission of phantoms for linearity, homogeneity, and long-term stability. A photon time-of-flight spectroscopy technique was used to characterize the optical properties of the phantoms. Results: In total, 24 phantoms classified into 4 types, namely homogeneous, multilayer, inclusion, and base phantoms, were manufactured. The phantoms exhibit linear behavior over the dosage range of UCNPs. The phantoms were found to be stable over a limited observed period of 4 months with a coefficient of variation of < 4 % . The deep tissue imaging case showed that increasing the thickness of tissue reduced the UCNP emission. Conclusions: A first-of-its-kind UCNP-based solid phantom recipe was developed, and four types of UCNP phantom tools to explore biophotonics applications were presented. The UCNP phantoms exhibited a linear behavior with dosage and were stable over time. An example case showed the potential use of the phantom for deep tissue imaging applications. With recent advance in the use of UCNPs for biophotonics, we believe our recipe and tools will play a pivotal role in the growth of the UCNPs for biophotonics applications.

Estimation of porcine pancreas optical properties in the 600-1100 nm wavelength range for light-based therapies.

This work reports the optical properties of porcine pancreatic tissue in the broad wavelength range of 600-1100 nm. Absorption and reduced scattering coefficients (µa and µs') of the ex vivo pancreas were obtained by means of Time-domain Diffuse Optical Spectroscopy. We have investigated different experimental conditions-including compression, repositioning, spatial sampling, temporal stability-the effect of the freezing procedure (fresh vs frozen-thawed pancreas), and finally inter-sample variability. Good repeatability under different experimental conditions was obtained (median coefficient of variation less than 8% and ~ 16% for µa and µs', respectively). Freezing-thawing the samples caused an irreversible threefold reduction of µs' and no effect on µa. The absorption and reduced scattering spectra averaged over different samples were in the range of 0.12-0.74 cm-1 and 12-21 cm-1 with an inter-sample variation of ~ 10% and ~ 40% for µa and µs', respectively. The calculated effective transport coefficient (µeff) for fresh pancreatic tissue shows that regions between 800-900 nm and 1050-1100 nm are similar and offer the lowest tissue attenuation in the considered range (i.e., µeff ranging from 2.4 to 2.7 cm-1). These data, describing specific light-pancreas interactions in the therapeutic optical window for the first time, provide pivotal information for planning of light-based thermotherapies (e.g., laser ablation) and instruction of light transport models for biophotonic applications involving this organ.

Multi-laboratory performance assessment of diffuse optics instruments: the BitMap exercise.

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.

Reproducibility of identical solid phantoms.

SIGNIFICANCE: Tissue-like solid phantoms with identical optical properties, known within tolerant uncertainty, are of crucial importance in diffuse optics for instrumentation assessment, interlaboratory comparison studies, industrial standards, and multicentric clinical trials. AIM: The reproducibility in fabrication of homogeneous solid phantoms is focused based on spectra measurements by instrument comparisons grounded on the time-resolved diffuse optics. APPROACH: Epoxy-resin and silicone phantoms are considered as matrices and both employ three different instruments for time-resolved diffuse spectroscopy within the spectral range of 540 to 1100 nm. In particular, we fabricated two batches of five phantoms each in epoxy resin and silicone. Then, we evaluated the intra- and interbatch variability with respect to the instrument precision, by considering the coefficient of variation (CV) of absorption and reduced scattering coefficients. RESULTS: We observed a similar precision for the three instruments, within 2% for repeated measurements on the same phantom. For epoxy-resin phantoms, the intra- and the interbatch variability reached the instrument precision limit, demonstrating a very good phantom reproducibility. For the silicone phantoms, we observed larger values for intra- and interbatch variability. In particular, at worst, for reduced scattering coefficient interbatch CV was about 5%. CONCLUSIONS: Results suggest that the fabrication of solid phantoms, especially considering epoxy-resin matrix, is highly reproducible, even if they come from different batch fabrications and are measured using different instruments.

Time-domain NIRS system based on supercontinuum light source and multi-wavelength detection: validation for tissue oxygenation studies.

We present and validate a multi-wavelength time-domain near-infrared spectroscopy (TD-NIRS) system that avoids switching wavelengths and instead exploits the full capability of a supercontinuum light source by emitting and acquiring signals for the whole chosen range of wavelengths. The system was designed for muscle and brain oxygenation monitoring in a clinical environment. A pulsed supercontinuum laser emits broadband light and each of two detection modules acquires the distributions of times of flight of photons (DTOFs) for 16 spectral channels (used width 12.5 nm / channel), providing a total of 32 DTOFs at up to 3 Hz. Two emitting fibers and two detection fiber bundles allow simultaneous measurements at two positions on the tissue or at two source-detector separations. Three established protocols (BIP, MEDPHOT, and nEUROPt) were used to quantitatively assess the system's performance, including linearity, coupling, accuracy, and depth sensitivity. Measurements were performed on 32 homogeneous phantoms and two inhomogeneous phantoms (solid and liquid). Furthermore, measurements on two blood-lipid phantoms with a varied amount of blood and Intralipid provide the strongest validation for accurate tissue oximetry. The retrieved hemoglobin concentrations and oxygen saturation match well with the reference values that were obtained using a commercially available NIRS system (OxiplexTS) and a blood gas analyzer (ABL90 FLEX), except a discrepancy occurs for the lowest amount of Intralipid. In-vivo measurements on the forearm of three healthy volunteers during arterial (250 mmHg) and venous (60 mmHg) cuff occlusions provide an example of tissue monitoring during the expected hemodynamic changes that follow previously well-described physiologies. All results, including quantitative parameters, can be compared to other systems that report similar tests. Overall, the presented TD-NIRS system has an exemplary performance evaluated with state-of-the-art performance assessment methods.

Optical signatures of radiofrequency ablation in biological tissues.

Accurate monitoring of treatment is crucial in minimally-invasive radiofrequency ablation in oncology and cardiovascular disease. We investigated alterations in optical properties of ex-vivo bovine tissues of the liver, heart, muscle, and brain, undergoing the treatment. Time-domain diffuse optical spectroscopy was used, which enabled us to disentangle and quantify absorption and reduced scattering spectra. In addition to the well-known global (1) decrease in absorption, and (2) increase in reduced scattering, we uncovered new features based on sensitive detection of spectral changes. These absorption spectrum features are: (3) emergence of a peak around 840 nm, (4) redshift of the 760 nm deoxyhemoglobin peak, and (5) blueshift of the 970 nm water peak. Treatment temperatures above 100 °C led to (6) increased absorption at shorter wavelengths, and (7) further decrease in reduced scattering. This optical behavior provides new insights into tissue response to thermal treatment and sets the stage for optical monitoring of radiofrequency ablation.

Non-invasive investigation of adipose tissue by time domain diffuse optical spectroscopy.

The human abdominal region is very heterogeneous and stratified with subcutaneous adipose tissue (SAT) being one of the primary layers. Monitoring this tissue is crucial for diagnostic purposes and to estimate the effects of interventions like caloric restriction or bariatric surgery. However, the layered nature of the abdomen poses a major problem in monitoring the SAT in a non-invasive way by diffuse optics. In this work, we examine the possibility of using multi-distance broadband time domain diffuse optical spectroscopy to assess the human abdomen non-invasively. Broadband absorption and reduced scattering spectra from 600 to 1100 nm were acquired at 1, 2 and 3 cm source-detector distances on ten healthy adult male volunteers, and then analyzed using a homogeneous model as an initial step to understand the origin of the detected signal and how tissue should be modeled to derive quantitative information. The results exhibit a clear influence of the layered nature on the estimated optical properties. Clearly, the underlying muscle makes a relevant contribution in the spectra measured at the largest source-detector distance for thinner subjects related to blood and water absorption. More unexpectedly, also the thin superficial skin layer yields a direct contamination, leading to higher water content and steeper reduced scattering spectra at the shortest distance, as confirmed also by simulations. In conclusion, provided that data analysis properly accounts for the complex tissue structure, diffuse optics may offer great potential for the continuous non-invasive monitoring of abdominal fat.

Optical characterization of porcine tissues from various organs in the 650-1100†nm range using time-domain diffuse spectroscopy.

We present a systematic characterization of the optical properties (µa and µs') of nine representative ex vivo porcine tissues over a broadband spectrum (650-1100 nm). We applied time-resolved diffuse optical spectroscopy measurements for recovering the optical properties of porcine tissues depicting a realistic representation of the tissue heterogeneity and morphology likely to be found in different ex vivo tissues. The results demonstrate a large spectral and inter-tissue variation of optical properties. The data can be exploited for planning or simulating ex vivo experiments with various biophotonics techniques, or even to construct artificial structures mimicking specific pathologies exploiting the wide assortment in optical properties.

Solid phantom recipe for diffuse optics in biophotonics applications: a step towards anatomically correct 3D tissue phantoms.

We present a tissue mimicking optical phantom recipe to create robust well tested solid phantoms. The recipe consists of black silicone pigment (absorber), silica microspheres (scatterer) and silicone rubber (SiliGlass, bulk material). The phantom recipe was characterized over a broadband spectrum (600-1100 nm) for a wide range of optical properties (absorption 0.1-1 cm-1, reduced scattering 5-25 cm-1) that are relevant to human organs. The results of linearity show a proper scaling of optical properties as well as the absence of coupling between the absorber and scatterer at different concentrations. A reproducibility of 4% among different preparations was obtained, with a similar grade of spatial homogeneity. Finally, a 3D non-scattering mock-up phantom of an infant torso made with the same recipe bulk material (SiliGlass) was presented to project the futuristic aspect of our work that is 3D printing human organs of biomedical relevance.