Recent advances in Doppler techniques have enabled high sensitivity imaging of biological flow to measure blood velocities and vascular perfusion. Here we compare spectrometer-based and wavelength-swept Doppler OCT implementations theoretically and experimentally, characterizing the lower and upper observable velocity limits in each configuration. We specifically characterize the washout limit for Doppler OCT, the velocity at which signal degradation results in loss of flow information, which is valid for both quantitative and qualitative flow imaging techniques. We also clearly differentiate the washout effect from the separate phenomenon of phase wrapping. We demonstrate that the maximum detectable Doppler velocity is determined by the fringe washout limit and not phase wrapping. Both theory and experimental results from phantom flow data and retinal blood flow data demonstrate the superiority of the swept-source technique for imaging vessels with high flow rates.

Dual emissive luminescence properties of solid-state difluoroboron β-diketonate-poly(lactic acid) (BF2bdk-PLA) materials have been utilized as biological oxygen sensors. Dyes with red-shifted absorption and emission are important for multiplexing and in vivo imaging, thus hydroxyl-functionalized dinaphthoylmethane initiators and dye-PLA conjugates BF2dnm(X)PLA (X = H, Br, I) with extended conjugation were synthesized. The luminescent materials show red-shifted absorbance (~435 nm) and fluorescence tunability by molecular weight. Fluorescence colors range from yellow (~530 nm) in 10 – 12 kDa polymers to green (~490 nm) in 20 – 30 kDa polymers. Room-temperature phosphorescence (RTP) and thermally activated delayed fluorescence (TADF) are present under a nitrogen atmosphere. For the iodine-substituted derivative, BF2dnm(I)PLA, clearly distinguishable fluorescence (green) and phosphorescence (orange) peaks are present, making it ideal for ratiometric oxygen-sensing and imaging. Bromide and hydrogen analogues with weaker relative phosphorescence intensities and longer phosphorescence lifetimes can be used as highly sensitive, concentration independent, lifetime-based oxygen sensors or for gated emission detection. BF2dnm(I)PLA nanoparticles were taken up by T41 mouse mammary cells and successfully demonstrated differences in vitro ratiometric measurement of oxygen.

Variance processing methods in Fourier domain optical coherence tomography (FD-OCT) have enabled depth-resolved visualization of the capillary beds in the retina due to the development of imaging systems capable of acquiring A-scan data in the 100 kHz regime. However, acquisition of volumetric variance data sets still requires several seconds of acquisition time, even with high speed systems. Movement of the subject during this time span is sufficient to corrupt visualization of the vasculature. We demonstrate a method to eliminate motion artifacts in speckle variance FD-OCT images of the retinal vasculature by creating a composite image from multiple volumes of data acquired sequentially. Slight changes in the orientation of the subject’s eye relative to the optical system between acquired volumes may result in non-rigid warping of the image. Thus, we use a B-spline based free form deformation method to automatically register variance images from multiple volumes to obtain a motion-free composite image of the retinal vessels. We extend this technique to automatically mosaic individual vascular images into a widefield image of the retinal vasculature.

Phase sensing implementations of spectral domain optical coherence tomography (SDOCT) have demonstrated the ability to measure nanometer-scale temporal and spatial profiles of samples. However, the phase information suffers from a 2pi ambiguity that limits observations of larger sample displacements to lengths less than half the source center wavelength. We introduce a synthetic wavelength phase unwrapping technique in SDOCT that uses spectral windowing and corrects the 2pi ambiguity, providing accurate measurements of sample motion with information gained from standard SDOCT processing. We demonstrate this technique by using a common path implementation of SDOCT and correctly measure phase profiles from a phantom phase object and human epithelial cheek cells which produce multiple wrapping artifacts. Using a synthetic wavelength for phase unwrapping could prove useful in Doppler or other phase based implementations of OCT.

We have combined hyperspectral imaging with spectral domain optical coherence tomography (SDOCT) to non-invasively image changes in hemoglobin saturation, blood flow, microvessel morphology and sheer rate on the vessel wall with tumor growth. Changes in these hemodynamic variables were measured over 24 hours in dorsal skin fold window chamber tumors. There was a strong correlation between volumetric flow and hemoglobin saturation (ρ = 0.89, p = 9 × 10−6, N = 15), and a moderate correlation between shear rate on the vessel wall and hemoglobin saturation (ρ = 0.56, p = 0.03, N=15).