Brillouin microscopy monitors rapid responses in subcellular compartments.

Measurements and imaging of the mechanical response of biological cells are critical for understanding the mechanisms of many diseases, and for fundamental studies of energy, signal and force transduction. The recent emergence of Brillouin microscopy as a powerful non-contact, label-free way to non-invasively and non-destructively assess local viscoelastic properties provides an opportunity to expand the scope of biomechanical research to the sub-cellular level. Brillouin spectroscopy has recently been validated through static measurements of cell viscoelastic properties, however, fast (sub-second) measurements of sub-cellular cytomechanical changes have yet to be reported.

Next-generation endoscopic probe for detection of esophageal dysplasia using combined OCT and angle-resolved low-coherence interferometry.

Angle-resolved low-coherence interferometry (a/LCI) is an optical technique that enables depth-specific measurements of nuclear morphology, with applications to detecting epithelial cancers in various organs. Previous a/LCI setups have been limited by costly fiber-optic components and large footprints. Here, we present a novel a/LCI instrument incorporating a channel for optical coherence tomography (OCT) to provide real-time image guidance.

Comparing the segmentation of quantitative phase images of neurons using convolutional neural networks trained on simulated and augmented imagery.

Significance: Quantitative phase imaging (QPI) can visualize cellular morphology and measure dry mass. Automated segmentation of QPI imagery is desirable for tracking neuron growth. Convolutional neural networks (CNNs) have provided state-of-the-art results for image segmentation.

Dynamic nitrogen vacancy magnetometry by single-shot optical streaking microscopy.

Nitrogen vacancy diamonds have emerged as sensitive solid-state magnetic field sensors capable of producing diffraction limited and sub-diffraction field images. Here, for the first time, to our knowledge, we extend those measurements to high-speed imaging, which can be readily applied to analyze currents and magnetic field dynamics in circuits on a microscopic scale. To overcome detector acquisition rate limitations, we designed an optical streaking nitrogen vacancy microscope to acquire two-dimensional spatiotemporal kymograms.

Rapid and precise tracking of water influx and efflux across cell membranes induced by a pulsed electric field.

Quantitative measurements of water content within a single cell are notoriously difficult. In this work, we introduce a single-shot optical method for tracking the intracellular water content, by mass and volume, of a single cell at video rate. We utilize quantitative phase imaging and a priori knowledge of a spherical cellular geometry, leveraging a two-component mixture model to compute the intracellular water content.

Comprehensive single-shot biophysical cytometry using simultaneous quantitative phase imaging and Brillouin spectroscopy.

Single-cell analysis, or cytometry, is a ubiquitous tool in the biomedical sciences. Whereas most cytometers use fluorescent probes to ascertain the presence or absence of targeted molecules, biophysical parameters such as the cell density, refractive index, and viscosity are difficult to obtain. In this work, we combine two complementary techniques-quantitative phase imaging and Brillouin spectroscopy-into a label-free image cytometry platform capable of measuring more than a dozen biophysical properties of individual cells simultaneously.

Esophageal OCT Imaging Using a Paddle Probe Externally Attached to Endoscope.

Background And Aims: Endoscopic surveillance of Barrett's esophagus (BE) by white light examination is insufficient to diagnose dysplastic change. In this work, we describe an optical imaging method to obtain high-resolution cross-sectional imaging using a paddle-shaped probe affixed to the endoscope tip.

Methods: We integrated Optical Coherence Tomography (OCT), an optical imaging method that produces cross-sectional images, into a paddle probe attached to video endoscope.

Deep imaging with 1.3 µm dual-axis optical coherence tomography and an enhanced depth of focus.

For many clinical applications, such as dermatology, optical coherence tomography (OCT) suffers from limited penetration depth due primarily to the highly scattering nature of biological tissues. Here, we present a novel implementation of dual-axis optical coherence tomography (DA-OCT) that offers improved depth penetration in skin imaging at 1.3 µm compared to conventional OCT.

Visualizing bleb mass dynamics in single cells using quantitative phase microscopy.

Understanding biological responses to directed energy (DE) is critical to ensure the safety of personnel within the Department of Defense. At the Air Force Research Laboratory, we have developed or adapted advanced optical imaging systems that quantify biophysical responses to DE. One notable cellular response to DE exposure is the formation of blebs, or semi-spherical protrusions of the plasma membrane in living cells.

Optical coherence tomography of small intestine allograft biopsies using a handheld surgical probe.

Significance: The current gold standard for monitoring small intestinal transplant (IT) rejection is endoscopic visual assessment and biopsy of suspicious lesions; however, these lesions are only superficially visualized by endoscopy. Invasive biopsies provide a coarse sampling of tissue health without depicting the true presence and extent of any pathology. Optical coherence tomography (OCT) presents a potential alternative approach with significant advantages over traditional white-light endoscopy.

Quantitative phase microscopy monitors subcellular dynamics in single cells exposed to nanosecond pulsed electric fields.

A substantial body of literature exists to study the dynamics of single cells exposed to short duration (<1 μs), high peak power (~1 MV/m) transient electric fields. Much of this research is limited to traditional fluorescence-based microscopy techniques, which introduce exogenous agents to the culture and are only sensitive to a single molecular target. Quantitative phase imaging (QPI) is a coherent imaging modality which uses optical path length as a label-free contrast mechanism, and has proven highly effective for the study of single-cell dynamics.

Spatial scanning of a sample with two-dimensional angle-resolved low-coherence interferometry for analysis of anisotropic scatterers.

Angle-resolved low-coherence interferometry (a/LCI) measures depth-resolved angular scattering for cell nuclear morphology analysis. 2D a/LCI, developed to collect across two scattering planes, is currently limited by the lack of spatial scanning. Here we demonstrate 2D a/LCI scanning across a three-dimensional volume using an image rotation scheme and a scanning mirror.

Reconstruction of angle-resolved backscattering through a multimode fiber for cell nuclei and particle size determination.

We demonstrate reconstruction of angle-resolved optical backscattering after transmission through a multimode fiber. Angle-resolved backscattering is an important tool for particle sizing, and has been developed as a diagnostic modality for detecting epithelial precancer. In this work, we fully characterized the transfer function of a multimode fiber using a plane-wave illumination basis across two dimensions.

Multimodal Coherent Imaging of Retinal Biomarkers of Alzheimer's Disease in a Mouse Model.

We acquired depth-resolved light scattering measurements from the retinas of triple transgenic Alzheimer's Disease (3xTg-AD) mice and wild type (WT) age-matched controls using co-registered angle-resolved low-coherence interferometry (a/LCI) and optical coherence tomography (OCT). Angle-resolved light scattering measurements were acquired from the nerve fiber layer, outer plexiform layer, and retinal pigmented epithelium using image guidance and segmented thicknesses provided by co-registered OCT B-scans. Analysis of the OCT images showed a statistically significant thinning of the nerve fiber layer in AD mouse retinas compared to WT controls.

Optical coherence tomography through a rigid borescope applied to quantification of articular cartilage thickness in a porcine knee model.

There exists an unmet need for an optical coherence tomography (OCT) delivery scheme that is simple, robust, and applicable to general surgical applications. To deliver the beam in a narrow form factor, optical borescopes present an attractive potential solution. We present a method for enabling endoscopic delivery of OCT using a handheld rigid borescope adapted to a low-cost OCT engine.

Shear Modulus Measurement by Quantitative Phase Imaging and Correlation with Atomic Force Microscopy.

Many approaches have been developed to characterize cell elasticity. Among these, atomic force microscopy (AFM) combined with modeling has been widely used to characterize cellular compliance. However, such approaches are often limited by the difficulties associated with using a specific instrument and by the complexity of analyzing the measured data.

Light scattering methods for tissue diagnosis.

Light scattering has become a common biomedical research tool, enabling diagnostic sensitivity to myriad tissue alterations associated with disease. Light-tissue interactions are particularly attractive for diagnostics due to the variety of contrast mechanisms that can be used, including spectral, angle-resolved, and Fourier-domain detection. Photonic diagnostic tools offer further benefit in that they are non-ionizing, non-invasive, and give real-time feedback.

Angular range, sampling and noise considerations for inverse light scattering analysis of nuclear morphology.

In recent years, significant work has been devoted to the use of angle-resolved elastic scattering for the extraction of nuclear morphology in tissue. By treating the nucleus as a Mie scattering object, techniques such as angle-resolved low-coherence interferometry (a/LCI) have demonstrated substantial success in identifying nuclear alterations associated with dysplasia. Because optical biopsies are inherently noninvasive, only a small, discretized portion of the 4π scattering field can be collected from tissue, limiting the amount of information available for diagnostic purposes.

Response to Comment on "Is the nuclear refractive index lower than cytoplasm? Validation of phase measurements and implications for light scattering technologies": A Comment on "How a phase image of a cell with nucleus refractive index smaller than that of the cytoplasm should look like?", e201800033.

Recently, Maxim A. Yurkin commented on our paper "Is the nuclear refractive index lower than cytoplasm? Validation of phase measurements and implications for light scattering technologies" as well as on a complementary study "Cell nuclei have lower refractive index and mass density than cytoplasm" from Schürmann et al. In his comment, Yurkin concluded that quantitative phase images of cells with nuclei that are less optically dense than the cytoplasm must exhibit a characteristic concavity, the absence of which is evidence against our conclusion of a less-dense nucleus.

Comparison of imaging fiber bundles for coherence-domain imaging.

Use of imaging fiber bundles for coherence-domain imaging has remained limited to date. In this work, we provide characterization of commercially available imaging bundles for coherence-domain imaging, by evaluating their modal structure for applicability to interferometric imaging. We further examine custom fabricated bundles developed in collaboration with a corporate partner for their ability to reduce interelement optical path length variability and cross talk between elements.

Revealing the glass transition in shape memory polymers using Brillouin spectroscopy.

Emerging medical devices which employ shape memory polymers (SMPs) require precise measurements of the glass transition temperature (T) to ensure highly controlled shape recovery kinetics. Conventional techniques like differential scanning calorimetry (DSC) and dynamic mechanical analysis (DMA) have limitations that prevent utilization for certain devices, including limited accuracy and the need for sacrificial samples. In this report, we employ an approach based on Brillouin spectroscopy to probe the glass transition of SMPs rapidly, remotely, and nondestructively.

Scanning system for angle-resolved low-coherence interferometry.

Angle-resolved low-coherence interferometry (a/LCI) detects precancer by enabling depth-resolved measurements of nuclear morphology in vivo. A significant limitation of a/LCI is the point-probe nature of the method, sampling <0.5  mm before probe relocation is necessary.

nsPEF-induced PIP depletion, PLC activity and actin cytoskeletal cortex remodeling are responsible for post-exposure cellular swelling and blebbing.

Cell swelling and blebbing has been commonly observed following nanosecond pulsed electric field (nsPEF) exposure. The hypothesized origin of these effects is nanoporation of the plasma membrane (PM) followed by transmembrane diffusion of extracellular fluid and disassembly of cortical actin structures. This investigation will provide evidence that shows passive movement of fluid into the cell through nanopores and increase of intracellular osmotic pressure are not solely responsible for this observed phenomena.

Is the nuclear refractive index lower than cytoplasm? Validation of phase measurements and implications for light scattering technologies.

The refractive index (RI) of biological materials is a fundamental parameter for the optical characterization of living systems. Numerous light scattering technologies are grounded in a quantitative knowledge of the refractive index at cellular and subcellular scales. Recent work in quantitative phase microscopy (QPM) has called into question the widely held assumption that the index of the cell nucleus is greater than that of the cytoplasm, a result which disagrees with much of the current literature.

Optical Phase Measurements of Disorder Strength Link Microstructure to Cell Stiffness.

There have been sustained efforts on the part of cell biologists to understand the mechanisms by which cells respond to mechanical stimuli. To this end, many rheological tools have been developed to characterize cellular stiffness. However, measurement of cellular viscoelastic properties has been limited in scope by the nature of most microrheological methods, which require direct mechanical contact, applied at the single-cell level.