Ultrasound-Aided Large-Scale Optoacoustic Microscopy for Volumetric Angiography and Oximetry

Abstract

Given its direct relationship to tissue metabolism and various pathological processes, 3D mapping of blood oxygen saturation (sO2) is essential for advancing our knowledge on oxygen delivery to tissues and evaluating therapeutic efficacy. Optoacoustic microscopy has enabled label-free estimation of sO2 values by exploiting the spectrally distinctive absorption of hemoglobin in its oxygenated and deoxygenated forms. However, quantitative 3D mapping of sO2 distribution over large heterogenous tissue regions is commonly hindered due to the strong spatial and spectral variability of the excitation light fluence. Herein, we capitalize on hybridization between pulse-echo ultrasound and large-scale spectroscopic optoacoustic microscopy readings to accurately delineate the tissue surface, achieve depth-resolved tissue layer segmentation, and comprehensively evaluate the main causes behind inaccurate sO2 estimations with optoacoustic microscopy. Compensation for wavelength-dependent light fluence variations due to relative reflectance and attenuation through multiple tissue layers is further shown to remove spectral noise and restore physiologically relevant sO2 values in the images recorded from the mouse ear and the dorsal murine skin. The ultrasound-aided large-scale optoacoustic microscopy (uLSOM) approach is thus expected to enhance applicability of optoacoustic microscopy for quantitative label-free imaging of tissue oxygenation and metabolism.