Journal of Biomedical Optics最新文献

Significance: The relationship between spatial offset and tissue sensing depth is not well understood in spatial offset Raman spectroscopy (SORS). Detection of the subsurface biochemical composition could improve clinical translation of SORS-based methods, including for lumpectomy margin characterization in breast cancer surgery.

Aim: We aimed at developing an experimental method to establish a relationship between spatial offset in SORS and sampling depth. The technique was developed using a custom hyperspectral line-scanning imaging system optimized for Raman spectroscopy detection.

Approach: Bilayer phantoms were produced with top and bottom layers made of material with different Raman spectroscopy signatures, i.e., poly(dimethylsiloxane) polymer (PDMS) and Nylon. The top layer of PDMS had different values of absorption and reduced elastic scattering coefficients, as well as a thickness up to $\sim 3\mathrm{mm}$ . A metric was used, called spectral angle mapper, that allowed for comparing SORS measurements with reference spectra of pure PDMS and Nylon. That metric was used to develop a technique predicting sensing depth for different values of spatial offset. A proof-of-concept study was performed to assess the performance of the method in biological tissue, demonstrating detectability of protein-rich tissue across layers of Intralipid and porcine fat to simulate the optical properties of human adipose tissue.

Results: A total of 60 optical phantoms with varying optical properties and top layer thicknesses were imaged and processed to estimate sampling depth as a function of spatial offset. The study demonstrated the detectability of the underlying Nylon layer across a PDMS layer up to 3 mm in thickness. Similarly, the detectability of protein-rich tissue was demonstrated across layers of Intralipid up to 3 mm thick and $<2\mathrm{mm}$ for porcine fat.

Conclusions: We showed the feasibility of using bilayer solid optical phantoms to create correlation curves between the optimal spatial offset for a desired probed depth given the optical properties of the top layer. The technique could facilitate the clinical translation of SORS measurements for tumor detection and margins assessment.

Significance: Accurate estimation of hydrogel phantom elasticity in 3D cell culture systems provides valuable insights into cellular responses to various mechanical stimuli. Although reverberant wave elastography has been applied to measure hydrogel elasticity in 3D cell cultures using multi-point loading, achieving a high-quality reverberant displacement field remains critical for accurate reverberant wave elastography.

Aim: We develop an innovative approach using 3D-printed randomly distributed scatterers to improve displacement field quality in reverberant wave elastography, inspired by scattering-coded architectured boundaries in object localization.

Approach: Numerical simulations were performed to analyze the reverberant displacement fields under various loading conditions. The results were compared to determine the optimal loading configuration to enhance the reverberation level of the displacement field. Subsequently, both numerical and experimental reverberant wave elastography were carried out to validate the elasticity measurement with 3D-printed randomly distributed scatterers.

Results: The comparison of reverberant displacement patterns under various loading conditions revealed that the displacement pattern under circular loading with 64 scatterers most closely approximated a diffuse wave field, exhibiting both spatial uniformity and directional isotropy. Numerical reverberant wave elastography was subsequently performed, successfully demonstrating its capability for elasticity measurements. Furthermore, the shear wave speeds obtained through optical coherence elastography showed good agreement with shear rheometry measurements.

Conclusions: The developed 3D-printed randomly distributed scatterers successfully enhanced the quality of the reverberant displacement field for reverberant wave elastography. Our approach presents a novel and promising tool for quantifying tissue elasticity in reverberant wave elastography applications.

The editorial introduces the JBO Special Section "Advances in Optical Elastography" for Volume 30 Issue 12.

Significance: Topical photodynamic therapy (PDT) with protoporphyrin IX (PpIX) converted from 5-aminolevulinic acid (ALA) is a well-established noninvasive method of treating skin conditions and lesions. During PDT, there can be response dynamics within the tissue that are affected by the light delivery, seen with fractionated delivery and in subcurative priming delivery. Fractionated light doses can considerably increase efficacy of 5-ALA PDT response.

Aim: We aim to examine the changes in physiological blood flow, tissue oxygenation, and PpIX concentration during and after light delivery in topical ALA-PDT in nude mouse skin.

Approach: We compared three schemes of light delivery for topical ALA-PDT in nude mice, including (1) full light delivery without fractionation, (2) two equal fractions (50% and 50%) of light separated by 2 h, and (3) a 5% light dose fractionation by 2 h prior to the main 95% light dose. Tissue oxygen imaging was assessed with the hypoxia signal from delayed fluorescence of PpIX itself within the tissue, as well as by confirmation with Oxyphor phosphorescence lifetime quenching imaging.

Results: The results of blood flow imaging and hypoxia imaging from PpIX and oxygen imaging with Oxyphor each showed evidence of increased capillary flow and tissue oxygenation after the initial 5% light dose, increased at the side of irradiation. This increased capillary flow and tissue oxygenation are presumably from vasodilation and local capillary flow increase. PpIX replenishment occurs during the intervening dark period after the initial light delivery.

Conclusion: These observations suggest that increasing oxygen and capillary flow combined with increased PpIX production together yield increased PDT efficiency, amplified by this initial light dose from a photodynamic optical priming event occurring 2 h prior to full PDT light delivery.

Significance: Current methods of measuring dosimetry for photodynamic therapy (PDT) have proven to be inadequate in their inability to provide accurate, real-time, and spatially resolved monitoring without interrupting the PDT treatment.

Aim: Our goal was to develop and validate a combined treatment and dosimetry system capable of monitoring implicit and explicit dosimetry in real time during non-contact PDT.

Approach: By employing both fluorescence imaging and spatial frequency domain imaging (SFDI), designed with low-cost, off-the-shelf components, the combined imaging system would be able to provide information on the spatial distributions of photosensitizer concentrations, tissue oxygenation, and delivered light dose, all while monitoring the photobleaching dynamics of the photosensitizer. Although the concept behind the combined system is not specific to any one photosensitizer, we focused on designing the system for the endogenous PDT of Gram-positive bacteria which utilizes coproporphyrin III as the photosensitizer.

Results: The overall performance of the system was assessed, with the accuracy, precision, and resolution of the SFDI-derived optical property maps being determined to fall within comparable ranges to other systems, despite the $1.0{\mathrm{mm}}^{-1}$ spatial frequency utilized for the shorter wavelengths. After validating the ability of the system to correct for tissue-like optical properties, and thus produce accurate quantitative fluorescence images, a preliminary assessment of antimicrobial PDT photobleaching dosimetry was performed, and high correlations were found between the fluorescence and PDT outcomes.

Conclusions: Overall, the developed imaging system showcases the potential to enable a more thorough analysis of PDT dosimetry and the impact of different variables on treatment outcomes.

Significance: Methicillin-resistant Staphylococcus aureus (MRSA) biofilm infections present a critical challenge in orthopedic trauma surgery and are notoriously resistant to systemic antibiotic therapy. Noninvasive, quantitative imaging methods are urgently needed to assess biofilm burden and therapeutic efficacy, especially for emerging photodynamic therapy (PDT) strategies.

Aim: We aim to establish a quantitative framework using a combined bioluminescence and optical coherence tomography (OCT) imaging approach to correlate bioluminescent signal with viable MRSA burden in both planktonic and biofilm states and to determine how biofilm density and structure influence this relationship.

Approach: Bioluminescent MRSA (SAP231-luxCDABE) was cultured in planktonic and biofilm forms using in vitro growth models in 24-well plates and custom macrofluidic devices, respectively. Bacteria bioluminescence intensity (BLI), counted colony-forming units (CFU), and OCT-based biofilm thickness measurements were collected to construct linear regression models to evaluate how well BLI alone, or combined with biofilm density (CFU/volume), predicts bacterial counts across culture conditions.

Results: Bioluminescence strongly correlated with CFU in planktonic cultures ( $R^2=0.98$ ). In biofilms, BLI per CFU decreased with density, indicating metabolic downregulation, and BLI alone was less reliable ( $R^2=0.59$ ). Incorporating biofilm density (CFU/volume) improved prediction ( $R^2=0.84$ ). A joint model for both states showed excellent fit ( $R^2=0.985$ ), but the biofilm versus planktonic group remained a significant factor ( $p=0.002$ ), revealing systematic differences. This highlights the need for a mixed-model approach that segments subvolumes by morphological features to improve accurate, generalizable CFU estimation across both growth states.

Conclusions: Bioluminescence alone underestimates bacterial burden in dense, metabolically suppressed MRSA biofilms. The combination of BLI with OCT-derived structural metrics enables accurate, nondestructive quantification of viable bacterial load. This approach provides a robust toolset for preclinical evaluation of antimicrobial therapies, particularly for optimizing PDT dosimetry and assessing biofilm response in translational infection models.

Significance: The stiffness and compliance of biological tissues are key properties that often change in the presence of pathology, yet current shear wave elastography approaches using optical coherence tomography (OCT) face limitations due to slow image acquisition, sensitivity to motion artifacts, and reliance on advanced hardware, hindering clinical translation.

Aim: The aim is to develop and validate a practical, high-speed method for three-dimensional shear wave imaging compatible with standard OCT systems and wave propagation variability.

Approach: We introduce a technique for the rapid, asynchronous acquisition of three-dimensional shear wave fields. Our technique operates at conventional acquisition rates and utilizes pairs of B-scans, similar to angiography scanning protocols. This approach significantly reduces motion sensitivity and enhances acquisition speed, even with much denser lateral sampling. In addition, we present a technique for estimating the shear wave number, termed directional phase gradient analysis. This method computes the phase gradient of the autocorrelation of the directionally-filtered, complex-valued shear wave and is robust across unidirectional, partially diffuse, and fully diffuse shear wave conditions.

Results: We validated the accuracy of our techniques through direct comparison with phase-locked, synchronous-mode imaging in benchtop experiments using tissue-mimicking phantoms. Furthermore, we demonstrated their robustness to variations in wave orientation, excitation amplitude, and diffusivity, as confirmed by repeated measurements on the same sample under diverse conditions.

Conclusions: Together, these methods may offer a more practical approach for shear wave imaging without requiring modifications to existing clinical phase-stable OCT systems.

Significance: Optical coherence elastography (OCE) is an emerging technique for mapping tissue mechanical properties into an image, known as an elastogram, with microscale resolution. Although system characterization phantoms are widely used in OCE development, there is a critical need for tissue-mimicking phantoms that can more accurately replicate the complex structural and mechanical properties of tissues, particularly for validating clinical applications, such as in breast cancer.

Aim: We aim to investigate the effects of tissue-like structures on elastogram formation in a controlled environment by developing and characterizing two types of breast tissue-mimicking phantoms, replicating invasive ductal carcinoma (IDC) morphology and the other mimicking breast ductal networks.

Approach: We present a comprehensive methodology for fabricating breast-mimicking phantoms using optical coherence tomography and ductography images to provide information on tissue structure. The method employs 3D-printed molds, casting different silicone materials for IDC-mimicking phantoms and implementing a dissolving mold technique to create duct-mimicking phantoms, which can be tested in both empty and fluid-filled states.

Results: The IDC-mimicking phantom successfully replicates structural features as small as $100\mu m$ , revealing complex mechanical behaviors at tissue interfaces, including strain concentrations where tissues of different stiffness interact. The duct-mimicking phantom demonstrates distinct mechanical responses between configurations, with hollow ducts creating sharp discontinuities at boundaries, whereas fluid-filled ducts exhibit more gradual transitions in mechanical properties.

Conclusions: Our methodology demonstrates the capability to fabricate breast tissue-mimicking phantoms that reproduce both the structural and mechanical properties of breast tissue, providing a controlled environment for investigating OCE performance and understanding how tissue architecture influences elastogram formation, particularly at interfaces among different tissue types.

Significance: Accurate and timely characterization of brain tumors remains a major challenge in neurosurgery. Current intraoperative guidance relies on preoperative imaging modalities such as magnetic resonance imaging, positron emission tomography, or computed tomography, which are essential for surgical planning but become less reliable during surgery due to brain shift. Furthermore, postoperative tumor classification depends on histopathology, which requires weeks and can delay treatment decisions. No existing tool offers real-time, label-free, and spatially resolved biomolecular information to support both intraoperative guidance and early tissue assessment.

Aim: We developed HyperProbe1.1 (HP1.1), a hyperspectral imaging system designed to acquire comprehensive molecular and metabolic information from brain tissue without the need for contrast agents or staining.

Approach: HP1.1 captures reflectance images across a broad range of narrow spectral bands, enabling spatial mapping of hemoglobin, cytochrome c oxidase, and oxygen saturation. In addition, ultraviolet-excited autofluorescence imaging provides information on metabolic cofactors - nicotinamide adenine dinucleotide and flavin adenine dinucleotide - relevant for tumor characterization. The system was validated using standardized phantoms and ex vivo glioma samples.

Results: HP1.1 demonstrated strong performance in detecting spectral features across phantoms and in distinguishing glioma tissues of different histological grades, enabling the generation of rapid and spatially resolved molecular contrast maps.

Conclusions: By providing label-free, high-content, and rapid biomolecular imaging, HP1.1 represents a powerful platform for noninvasive tissue assessment in controlled experimental settings and paves the way for future intraoperative applications.

Significance: Optical coherence elastography (OCE) is a noninvasive imaging technique with high sensitivity and resolution that can be used for mucocutaneous imaging. Oral submucous fibrosis (OSF) is a chronic disease that has a tendency to become cancerous. Nevertheless, there are a few noninvasive methods for early detection of OSF.

Aim: A piezoelectric transducer-based (PZT) OCE technique was devised to noninvasively assess the structural and mechanical properties of mucosa in healthy and fibrotic oral diseases.

Approach: We first validated the accuracy and reliability of the OCE system for tissue elasticity detection by means of a heterogeneous agar model. The structural and biomechanical characteristics of the regional tissues were then evaluated by examining the oral mucosa of both healthy and fibrotic SD rats.

Results: Normal and fibrotic tissue stiffness differed significantly ( $p<0.05$ ). The elastic wave velocity was $6.44\pm 0.30m/s$ in the normal group and $14.2\pm 0.91m/s$ in the fibrotic group. After converting the results to Young's modulus, the stiffness of the healthy buccal tissues and the fibrotic buccal tissues were $130.71\pm 12.01$ and $636.15\pm 79.17\mathrm{kPa}$ , respectively ( $p<0.05$ ).

Conclusions: OCE can differentiate between normal and fibrotic tissue based on elasticity and optical properties. Healthy buccal tissues were softer than diseased tissues.