Spectral photoacoustic imaging in combination with unmixing techniques may be applied to retrieve information about high-risk features present in atherosclerotic plaques, possibly providing prognostic insights into future stroke events. We present the photoacoustic spectral contrast found in 12 systematically scanned advanced atherosclerotic plaques in the near-infrared wavelength range (850–1250 nm). The main absorbers are lipid, water, and hemoglobin, with the highest photoacoustic intensities at the lipid's second overtone at 1190 and 1210 nm. Linear unmixing resulted in visualizing regions with high lipid and hemoglobin absorption, corresponding to the histological presence of lipid and intraplaque hemorrhage. A non-negative matrix factorization approach reveals differences in lipid spectral contrast, providing potential insights into the vulnerability of atherosclerotic plaque. These results provide a reference for future, more complex, in vivo photoacoustic imaging of carotid artery atherosclerosis, potentially contributing to assessing the risk of future events and treatment decision.

Photothermal optical coherence microscopy (PT-OCM) combines the high-resolution, label-free morphological imaging of OCM with the ability to discriminate tissue composition through phase-sensitive photothermal imaging. In this study, we perform 2D imaging of human carotid endarterectomies to spectrally determine lipid distribution, with verification via histologically stained samples. The structural information from OCM is combined with the spectral information gained from measuring the resulting sample surface displacement from thermoelastic expansion, following light irradiation. PT-OCM is thus demonstrated as a potential tool in the investigation of atherosclerotic plaque lipids, contributing towards the understanding of plaque instability.