Photon absorption remote sensing (PARS): comprehensive absorption imaging enabling label-free biomolecule characterization and mapping

Abstract

Label-free optical absorption microscopy techniques continue to evolve as promising tools for label-free histopathological imaging of cells and tissues. However, critical challenges relating to specificity and contrast, as compared to current gold-standard methods continue to hamper adoption. This work introduces Photon Absorption Remote Sensing (PARS), a new absorption microscope modality, which simultaneously captures the dominant de-excitation processes following an absorption event. In PARS, radiative (auto-fluorescence) and non-radiative (photothermal and photoacoustic) relaxation processes are collected simultaneously, providing enhanced specificity to a range of biomolecules. As an example, a multiwavelength PARS system featuring UV (266 nm) and visible (532 nm) excitation is applied to imaging human skin, and murine brain tissue samples. It is shown that PARS can directly characterize, differentiate, and unmix, clinically relevant biomolecules inside complex tissues samples using established statistical processing methods. Gaussian mixture models (GMM) are used to characterize clinically relevant biomolecules (e.g., white, and gray matter) based on their PARS signals, while non-negative least squares (NNLS) is applied to map the biomolecule abundance in murine brain tissues, without stained ground truth images or deep-learning methods. PARS unmixing and abundance estimates are directly validated and compared against chemically stained ground truth images, and deep learning based-image transforms. Overall, it is found that the PARS unique and rich contrast may provide comprehensive, and otherwise inaccessible, label-free characterization of molecular pathology, representing a new source of data to develop AI and machine learning methods for diagnostics and visualization.