Scattering oblique plane microscopy for in-vivo blood cell imaging.

Oblique plane microscopy (OPM) enables high speed, volumetric fluorescence imaging through a single-objective geometry. While these advantages have positioned OPM as a valuable tool to probe biological questions in animal models, its potential for in vivo human imaging is largely unexplored due to its typical use with exogenous fluorescent dyes. Here we introduce a scattering-contrast oblique plane microscope (sOPM) and demonstrate label-free imaging of blood cells flowing through human capillaries in vivo. The sOPM illuminates a capillary bed in the ventral tongue with an oblique light sheet, and images side- and back- scattered signal from blood cells. By synchronizing the sOPM with a conventional capillaroscope, we acquire paired widefield and axial images of blood cells flowing through a capillary loop. The widefield capillaroscope image provides absorption contrast and confirms the presence of red blood cells (RBCs), while the sOPM image may aid in determining whether optical absorption gaps (OAGs) between RBCs have cellular or acellular composition. Further, we demonstrate consequential differences between fluorescence and scattering versions of OPM by imaging the same polystyrene beads sequentially with each technique. Lastly, we substantiate in vivo observations by imaging isolated red blood cells, white blood cells, and platelets in vitro using 3D agar phantoms. These results demonstrate a promising new avenue towards in vivo blood analysis.

Calibration of liquid crystal variable retarders using a common-path interferometer and fit of a closed-form expression for the retardance curve.

A liquid crystal variable retarder (LCVR) enables fast, automated control of retardance that can be used as a variable waveplate in polarimetric instruments. However, precise control of the polarization state requires calibration of the LCVR. A manufacturer calibration curve is typically supplied for a single specific wavelength and temperature, but for applications under different conditions, additional calibration is needed. Calibration is typically performed with crossed polarizers to generate an intensity curve that is converted to retardance, but this method is prone to noise when retardance is close to zero. Here, we demonstrate a simple common-path Sagnac interferometer to measure retardance and provide open source software for automated generation of calibration curves for retardance as a function of wavelength and voltage. We also provide a curve fitting method and closed-form functional representation that outputs the voltage needed to achieve a desired retardance given a specified wavelength.

Platform for quantitative multiscale imaging of tissue composition.

Changes in the multi-level physical structure of biological features going from cellular to tissue level composition is a key factor in many major diseases. However, we are only beginning to understand the role of these structural changes because there are few dedicated multiscale imaging platforms with sensitivity at both the cellular and macrostructural spatial scale. A single platform reduces bias and complications from multiple sample preparation methods and can ease image registration. In order to address these needs, we have developed a multiscale imaging system using a range of imaging modalities sensitive to tissue composition: Ultrasound, Second Harmonic Generation Microscopy, Multiphoton Microscopy, Optical Coherence Tomography, and Enhanced Backscattering. This paper details the system design, the calibration for each modality, and a demonstration experiment imaging a rabbit eye.

Noise reduction in supercontinuum sources for OCT by single-pulse spectral normalization.

Supercontinuum (SC) sources offer high illumination power from a single mode fiber with large spectral bandwidth including the visible spectrum, a growing application area for Optical Coherence Tomography (OCT). However, SC spectra suffer from pulse-to-pulse variations, increasing noise in the resulting images. By simultaneously collecting a normalization spectrum, OCT image noise can be reduced by more than half (7 dB) for single pulses without any pulse averaging using only simple optical components.

Quantifying optical properties with visible and near-infrared optical coherence tomography to visualize esophageal microwave ablation zones.

Microwave ablation is a minimally invasive image guided thermal therapy for cancer that can be adapted to endoscope use in the gastrointestinal (GI) tract. Microwave ablation in the GI tract requires precise control over the ablation zone that could be guided by high resolution imaging with quantitative contrast. Optical coherence tomography (OCT) provides ideal imaging resolution and allows for the quantification of tissue scattering properties to characterize ablated tissue. Visible and near-infrared OCT image analysis demonstrated increased scattering coefficients (µs ) in ablated versus normal tissues (Vis: 347.8%, NIR: 415.0%) and shows the potential for both wavelength ranges to provide quantitative contrast. These data suggest OCT could provide quantitative image guidance and valuable information about antenna performance in vivo.