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Background Breast density is a strong and potentially modifiable breast cancer risk factor. Almost everything we know about breast density has been derived from mammography, and therefore, very little is known about breast density in younger women aged Methods Breast tissue composition (percent water, collagen, and lipid content) was measured in 539 women aged 18–40 using OBS. For a subset of 169 women, breast density was also measured via DXA (percent fibroglandular dense volume (%FGV), absolute dense volume (FGV), and non-dense volume (NFGV)). Acceptability of the measurement procedures was assessed using an adapted validated questionnaire. Performance was assessed by examining the correlation and agreement between the measures and their associations with known determinants of mammographic breast density. Results Over 93% of participants deemed OBS and DXA to be acceptable. The correlation between OBS-%water + collagen and %FGV was 0.48. Age and BMI were inversely associated with OBS-%water + collagen and %FGV and positively associated with OBS-%lipid and NFGV. Conclusions OBS and DXA provide acceptable and viable alternative methods to measure breast density in younger women aged 18–40 years.

Supplementary Figure 6 from Imaging of Human Lymph Nodes Using Optical Coherence Tomography: Potential for Staging Cancer

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Supplementary Figure 3 from Imaging of Human Lymph Nodes Using Optical Coherence Tomography: Potential for Staging Cancer

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Supplementary Figure 4 from Imaging of Human Lymph Nodes Using Optical Coherence Tomography: Potential for Staging Cancer

Supplementary Figure 9 from Imaging of Human Lymph Nodes Using Optical Coherence Tomography: Potential for Staging Cancer

Supplementary Figure 2 from Imaging of Human Lymph Nodes Using Optical Coherence Tomography: Potential for Staging Cancer

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Supplementary Figure 5 from Imaging of Human Lymph Nodes Using Optical Coherence Tomography: Potential for Staging Cancer

Mechanisms of tissue deformation in OCME

T2DM individuals exhibit macro- and microvascular dysfunction that is pronounced in the legs. Cocoa acutely increases endothelial function of the brachial and the femoral arteries and dilation of microvasculature in the feet of T2DM and healthy individuals.

Imaging needles consist of highly miniaturized focusing optics encased within a hypodermic needle. The needles may be inserted tens of millimeters into tissue and have the potential to visualize diseased cells well beyond the penetration depth of optical techniques applied externally. Multimodal imaging needles acquire multiple types of optical signals to differentiate cell types. However, their use has not previously been demonstrated with live cells.We demonstrate the ability of a multimodal imaging needle to differentiate cell types through simultaneous optical coherence tomography (OCT) and fluorescence imaging.We characterize the performance of a multimodal imaging needle. This is paired with a fluorescent analog of the therapeutic drug, tamoxifen, which enables cell-specific fluorescent labeling of estrogen receptor-positive (ER+) breast cancer cells. We perform simultaneous OCT and fluorescence in situ imaging on MCF-7 ER+ breast cancer cells and MDA-MB-231 ER- cells. Images are compared against unlabeled control samples and correlated with standard confocal microscopy images.We establish the feasibility of imaging live cells with these miniaturized imaging probes by showing clear differentiation between cancerous cells.Imaging needles have the potential to aid in the detection of specific cancer cells within solid tissue.

Optical coherence tomography (OCT) is a non-contact method for imaging the topological and internal microstructure of samples in three dimensions. OCT can be configured as a conventional microscope, as an ophthalmic scanner, or using endoscopes and small diameter catheters for accessing internal biological organs. In this Primer, we describe the principles underpinning the different instrument configurations that are tailored to distinct imaging applications and explain the origin of signal, based on light scattering and propagation. Although OCT has been used for imaging inanimate objects, we focus our discussion on biological and medical imaging. We examine the signal processing methods and algorithms that make OCT exquisitely sensitive to reflections as weak as just a few photons and that reveal functional information in addition to structure. Image processing, display and interpretation, which are all critical for effective biomedical imaging, are discussed in the context of specific applications. Finally, we consider image artifacts and limitations that commonly arise and reflect on future advances and opportunities.

The visualization and assessment of retinal microvasculature are important in the study, diagnosis, monitoring, and guidance of treatment of ocular and systemic diseases. With the introduction of optical coherence tomography angiography (OCTA), it has become possible to visualize the retinal microvasculature volumetrically and without a contrast agent. Many lab-based and commercial clinical instruments, imaging protocols and data analysis methods and metrics, have been applied, often inconsistently, resulting in a confusing picture that represents a major barrier to progress in applying OCTA to reduce the burden of disease. Open data and software sharing, and cross-comparison and pooling of data from different studies are rare. These inabilities have impeded building the large databases of annotated OCTA images of healthy and diseased retinas that are necessary to study and define characteristics of specific conditions. This paper addresses the steps needed to standardize OCTA imaging of the human retina to address these limitations. Through review of the OCTA literature, we identify issues and inconsistencies and propose minimum standards for imaging protocols, data analysis methods, metrics, reporting of findings, and clinical practice and, where this is not possible, we identify areas that require further investigation. We hope that this paper will encourage the unification of imaging protocols in OCTA, promote transparency in the process of data collection, analysis, and reporting, and facilitate increasing the impact of OCTA on retinal healthcare delivery and life science investigations.

Purpose Microperimetry measures differential light sensitivity (DLS) at specific retinal locations. The aim of this study is to examine the variation in DLS across the macula and the contribution to this variation of cone distribution metrics and retinal eccentricity. Methods Forty healthy eyes of 40 subjects were examined by microperimetry (MAIA) and adaptive optics imaging (rtx1). Retinal DLS was measured using the grid patterns: foveal (2°-3°), macular (3°-7°), and meridional (2°-8° on horizontal and vertical meridians). Cone density (CD), distribution regularity, and intercone distance (ICD) were calculated at the respective test loci coordinates. Linear mixed-effects regression was used to examine the association between cone distribution metrics and loci eccentricity, and retinal DLS. Results An eccentricity-dependent reduction in DLS was observed on all MAIA grids, which was greatest at the foveal-parafoveal junction (2°-3°) (-0.58 dB per degree, 95% confidence interval [CI]; -0.91 to -0.24 dB, P < 0.01). Retinal DLS across the meridional grid changed significantly with each 1000 cells/deg2 change in CD (0.85 dB, 95% CI; 0.10 to 1.61 dB, P = 0.03), but not with each arcmin change in ICD (1.36 dB, 95% CI; -2.93 to 0.20 dB, P = 0.09). Conclusions We demonstrate significant variation in DLS across the macula. Topographical change in cone separation is an important determinant of the variation in DLS at the foveal-parafoveal junction. We caution the extrapolation of changes in DLS measurements to cone distribution because the relationship between these variables is complex. Translational Relevance Cone density is an independent determinant of DLS in the foveal-parafoveal junction in healthy eyes.

Polarization-sensitive optical coherence tomography (PS-OCT) measures the depth-dependent polarization states of light backscattered in tissue. We have developed algorithms for the robust reconstruction of the local, depth-resolved OAx orientation, which recursively compensate for the cumulative effect of the preceding tissue layers. Local OAx imaging in healthy human skin in vivo revealed dense, weaving patterns that are imperceptible in OCT intensity tomograms and that suggest a mesh-like tissue organization, consistent with the morphology of dermal collagen. Local OAx orientation as a contrast mechanism merits further exploration for applications in dermatology and in other tissues and organs presenting intrinsic birefringence contrast.

Mammographic breast density is a strong breast cancer risk factor, and its routine clinical measurement could potentially be used to identify women at higher risk of breast cancer and/or monitor primary prevention strategies. Previous reports of optical breast spectroscopy (OBS), a novel approach to measuring breast density, demonstrated that it is safe (no ionizing radiation), portable, low-cost, and does not require image interpretation but have been limited to small, single-center studies. Reference measurements taken on a phantom breast prior to and after each woman’s OBS assessment are required for the calibration of the system transfer function as a part of processing participant data. To inform the validity of participant data, a detailed description of the reference measurements and a repeatability analysis of these measurements taken before and after participant assessment is presented. Reference measurements for OBS from 539 women aged 18–40 years were obtained as a part of a high-throughput epidemiological pilot study. Of these, measurements from 20 women with no useable data due to device failure (3.7%) were excluded and from another 12 women due to user error. The intra-class correlation (ICC) within complete pairs of reference data (taken before and after assessment) was high (all ICC > 0.84). The analysis presented here confirms the OBS participant data as valid for use in ongoing epidemiological research, providing further supporting evidence of OBS as a measure of breast density. A novel method of measuring breast density is needed to bridge large gaps in the knowledge of breast density in younger women and its relation to later-life breast cancer risk.

Significance: To advance our understanding of the contrast observed when imaging with polarization-sensitive optical coherence tomography (PS-OCT) and its correlation with oral cancerous pathologies, a detailed comparison with histology provided via ex vivo fixed tissue is required. The effects of tissue fixation, however, on such polarization-based contrast have not yet been investigated. Aim: A study was performed to assess the impact of tissue fixation on depth-resolved (i.e., local) birefringence measured with PS-OCT. Approach: A PS-OCT system based on depth-encoded polarization multiplexing and polarization-diverse detection was used to measure the Jones matrix of a sample. A wide variety of ex vivo samples were measured freshly after excision and 24 h after fixation, consistent with standard pathology. Some samples were also measured 48 h after fixation. Results: The tissue fixation does not diminish the birefringence contrast. Statistically significant changes were observed in 11 out of 12 samples; these changes represented an increase in contrast, overall, by 11% on average. Conclusions: We conclude that the fixed samples are suitable for studies seeking a deeper understanding of birefringence contrast in oral tissue pathology. The enhancement of contrast removes the need to image immediately postexcision and will facilitate future investigations with PS-OCT and other advanced polarization-sensitive microscopy methods, such as mapping of the local optic axis with PS-OCT and PS-optical coherence microscopy.

We show that polarization-sensitive optical coherence tomography angiography (PS-OCTA) based on full Jones matrix assessment of speckle decorrelation offers improved contrast and depth of vessel imaging over conventional OCTA. We determine how best to combine the individual Jones matrix elements and compare the resulting image quality to that of a conventional OCT scanner by co-locating and imaging the same skin locations with closely matched scanning setups. Vessel projection images from finger and forearm skin demonstrate the benefits of Jones matrix-based PS-OCTA. Our study provides a promising starting point and a useful reference for future pre-clinical and clinical applications of Jones matrix-based PS-OCTA.

Significance: Pulsatility is a vital characteristic of the cardiovascular system. Characterization of the pulsatility pattern locally in the peripheral microvasculature is currently not readily available and would provide an additional source of information which may prove important in understanding the pathophysiology of arterial stiffening, vascular ageing, and their linkage with cardiovascular disease development. Aim: We aim to confirm the suitability of speckle decorrelation optical coherence tomography angiography (OCTA) under various non-contact/contact scanning protocols for the visualization of pulsatility patterns in vessel-free tissue and in the microvasculature of peripheral human skin. Results: Results from 5 healthy subjects show distinct pulsatile patterns both in vessel-free tissue with either non-contact or contact imaging and in individual microvessels with contact imaging; respectively, likely caused by the pulsatile pressure and pulsatile blood flow. The pulse rates show good agreement with those from pulse oximetry, confirming that the pulsatile signatures reflect pulsatile hemodynamics. Conclusions: This study demonstrates the potential of speckle decorrelation OCTA for measuring localized peripheral cutaneous pulsatility and defines scanning protocols necessary to undertake such measurements. Non-contact imaging should be used for the study of pulsatility in vessel-free tissue and contact imaging with strong mechanical coupling in individual microvessels. Further studies of microcirculation based upon this method and protocols are warranted.

This chapter describes the main technology and methods of optical elastography—a suite of techniques that use optical methods to form images on the microscale of the mechanical properties of tissues. Optical elastography has recently shown compelling potential for the microscale characterization of soft tissues and cellular mechanics. Current developments are described in the context of their application, placed in perspective by touching briefly upon the history and likely future directions of the field. This chapter is mainly focused on two optical elastography techniques that have become prominent over the past 5 years: optical coherence elastography and Brillouin microscopy; however, other techniques, such as laser speckle imaging, photoacoustic elastography, and ultrasound-modulated optical tomography are briefly discussed in the context of tissue elasticity imaging.

High extracellular matrix (ECM) content in solid cancers impairs tumour perfusion and thus access of imaging and therapeutic agents. We have devised a new approach to degrade tumour ECM, which improves uptake of circulating compounds. We target the immune‐modulating cytokine, tumour necrosis factor alpha (TNFα), to tumours using a newly discovered peptide ligand referred to as CSG. This peptide binds to laminin–nidogen complexes in the ECM of mouse and human carcinomas with little or no peptide detected in normal tissues, and it selectively delivers a recombinant TNFα‐CSG fusion protein to tumour ECM in tumour‐bearing mice. Intravenously injected TNFα‐CSG triggered robust immune cell infiltration in mouse tumours, particularly in the ECM‐rich zones. The immune cell influx was accompanied by extensive ECM degradation, reduction in tumour stiffness, dilation of tumour blood vessels, improved perfusion and greater intratumoral uptake of the contrast agents gadoteridol and iron oxide nanoparticles. Suppressed tumour growth and prolonged survival of tumour‐bearing mice were observed. These effects were attainable without the usually severe toxic side effects of TNFα., This study establishes a new approach to treat solid tumours by immune modulation and ECM depletion. The developed agent, TNFα‐CSG, has dual capacity as an immunotherapeutic and ECM reducing agent which improves tumour perfusion and drug delivery.

Cellular-scale imaging of the mechanical properties of tissue has helped to reveal the origins of disease; however, cellular-scale resolution is not readily achievable in intact tissue volumes. Here, we demonstrate volumetric imaging of Young’s modulus using ultrahigh-resolution optical coherence elastography, and apply it to characterizing the stiffness of mouse aortas. We achieve isotropic resolution of better than 15 μm over a 1-mm lateral field of view through the entire depth of an intact aortic wall. We employ a method of quasi-static compression elastography that measures volumetric axial strain and uses a compliant, transparent layer to measure surface axial stress. This combination is used to estimate Young’s modulus throughout the volume. We demonstrate differentiation by stiffness of individual elastic lamellae and vascular smooth muscle. We observe stiffening of the aorta in regulator of G protein signaling 5-deficient mice, a model that is linked to vascular remodeling and fibrosis. We observe increased stiffness with proximity to the heart, as well as regions with micro-structural and micro-mechanical signatures characteristic of fibrous and lipid-rich tissue. High-resolution imaging of Young’s modulus with optical coherence elastography may become an important tool in vascular biology and in other fields concerned with understanding the role of mechanics within the complex three-dimensional architecture of tissue.

Many bioimaging studies, including those in engineered tissue constructs, intravital microscopy in animal models, and medical imaging in humans, require cellular-resolution imaging of structures deep within a sample. Yet, many of the current approaches are limited in terms of resolution, but also in invasiveness, repeatable imaging of the same location, and accessible imaging depth. We coin the term micro-endomicroscope to describe the emerging class of small, cellular-resolution endoscopic imaging systems designed to image cells in situ while minimizing perturbation of the sample. In this Perspective, we motivate the need for further development of micro-endomicroscopes, highlighting applications that would greatly benefit, reviewing progress, and considering how photonics might contribute. We identify areas ripe for technological development, such as micro-scanners and small lens systems, that would advance micro-endomicroscope performance. With the right developments in photonics, many possibilities exist for new minimally invasive translatable imaging tools across the scientific, pre-clinical, and clinical spectrum: from longitudinal studies of engineered tissue constructs, to tracking disease progression in animal models, to expanding the ability to diagnose and develop treatments for diseases without the need for invasive medical procedures.

Optical elastography, the use of optics to characterize and map the mechanical properties of biological tissue, involves measuring the deformation of tissue in response to a load. Such measurements may be used to form an image of a mechanical property, often elastic modulus, with the resulting mechanical contrast complementary to the more familiar optical contrast. Optical elastography is experiencing new impetus in response to developments in the closely related fields of cell mechanics and medical imaging, aided by advances in photonics technology, and through probing the microscale between that of cells and whole tissues. Two techniques — optical coherence elastography and Brillouin microscopy — have recently shown particular promise for medical applications, such as in ophthalmology and oncology, and as new techniques in cell mechanics.

This feature issue of Biomedical Optics Express represents a cross-section of the most recent work in tissue optics, including exciting developments in tissue optical clearing, deep tissue imaging, optical elastography, nanophotonics in tissue, and therapeutic applications of light, amongst others. A collection of 33 papers provides a comprehensive overview of current research in tissue optics, much of it inspired and informed by the pioneering work of Prof. Valery Tuchin. The issue contains three invited manuscripts and several mini-reviews that we hope will benefit researchers in this exciting area.

Non-Gaussian beams can provide extended depth of focus (DOF) at constant and potentially uncompromised transverse resolution, as well as a degree of self-reconstruction after beam shadowing, which may be present in tissue imaging. Hence such beams are being developed for imaging systems throughout many disciplines, including endoscopic imaging, where they hold great potential. General possibilities include up to more than 20-fold extension of DOF, tunable working distance, imaging around obstacles and integrated all-fiber designs. In all-fiber based optical imaging systems; however, these advantages are limited by system design considerations. Trade-offs between miniaturization, extended DOF, SNR, and fiber availability arise, and estimating the effects of design modifications can be difficult and time consuming. We model zero-order quasi-Bessel illumination and detection for a range of common probe and sample materials based on an analytic solution of the Fresnel diffraction integral and compare the results to Gaussian beams. We show that these beams, on scales that match optical fiber dimensions, generally have an upper limit for the spot size above which their distinct advantages over Gaussian beams fade. Similarly, we show the existence of a lower limit of practical performance of quasi-Bessel beams, where the imaging SNR penalty compared to a Gaussian beam becomes significant. Additionally to general theoretic considerations we discuss designs, modeling and characterization of all-fiber imaging probes. This work provides an accessible overview for researchers to estimate what potential benefit non-Gaussian beams can introduce into their optical imaging system.

We present a new optical coherence tomography (OCT) angiography method for imaging tissue microvasculature in vivo based on the characteristic frequency-domain flow signature in a short time series of a single voxel. The angiography signal is generated by Fourier transforming the OCT signal time series from a given voxel in multiple acquisitions and computing the average magnitude of non-zero (high-pass) frequency components. Larger temporal variations of the OCT signal caused by blood flow result in higher values of the average magnitude in the frequency domain compared to those from static tissue. Weighting of the signal by the inverse of the zero-frequency component (i.e., the sum of the OCT signal time series) improves vessel contrast in flow regions of low OCT signal. The method is demonstrated on a fabricated flow phantom and on human skin in vivo and, at only 5 time points per voxel, shows enhanced vessel contrast in comparison to conventional correlation mapping/speckle decorrelation and speckle variance methods.

The guest editors introduce a feature issue containing papers based on research presented at the OSA Biophotonics Congress (the former BIOMED) 20-23 April 2020, in the first all virtual, web conference format undertaken by OSA.

We demonstrate the use of the near-infrared attenuation coefficient, measured using optical coherence tomography (OCT), in longitudinal assessment of hypertrophic burn scars undergoing fractional laser treatment. The measurement method incorporates blood vessel detection by speckle decorrelation and masking, and a robust regression estimator to produce 2D en face parametric images of the attenuation coefficient of the dermis. Through reliable co-location of the field of view across pre- and post-treatment imaging sessions, the study was able to quantify changes in the attenuation coefficient of the dermis over a period of ~20 weeks in seven patients. Minimal variation was observed in the mean attenuation coefficient of normal skin and control (untreated) mature scars, as expected. However, a significant decrease (13 ± 5%, mean ± standard deviation) was observed in the treated mature scars, resulting in a greater distinction from normal skin in response to localized damage from the laser treatment. By contrast, we observed an increase in the mean attenuation coefficient of treated (31 ± 27%) and control (27 ± 20%) immature scars, with numerical values incrementally approaching normal skin as the healing progressed. This pilot study supports conducting a more extensive investigation of OCT attenuation imaging for quantitative longitudinal monitoring of scars. (Figure presented.) En face 2D OCT attenuation coefficient map of a treated immature scar derived from the pre-treatment (top) and the post-treatment (bottom) scans. (Vasculature (black) is masked out.) The scale bars are 0.5 mm.