Multiscale Vascular Enhancement Filter Applied to <italic>In Vivo</italic> Morphologic and Functional Photoacoustic Imaging of Rat Ocular Vasculature

Optical-resolution photoacoustic microscopy (OR-PAM) is used for <italic>in vivo</italic> imaging of a variety of albino and pigmented eyes taking advantages of requiring no exogenous dye, performing high-resolution imaging, and achieving morphologic and functional imaging at the same time. However, to accurately diagnose the ophthalmic disease in the OR-PAM images, vascular enhancement algorithms are necessary for extracting vessels and quantifying them correctly. Vascular enhancement algorithms developed for other imaging technologies, are not suitable to be used for OR-PAM, because of the underlying differences in the physics of the formation of images. In this study, a new vascular enhancement algorithm called photoacoustic imaging vasculature enhancement filter (PAIVEF) is proposed, which not only enhances vasculature including micro-vessels signals, suppresses noise signals effectively, but also achieves highly sensitive and accurate enhancement of the vasculature within a large depth range in and out of the system&#x0027;s depth of focus (DOF). Using the PAIVEF, the morphologic and functional 3D images of the whole rat&#x0027;s ocular anterior vasculature segment was displayed simultaneously for a depth range of &#x223C;0.6 mm, which was &#x223C;7 times of the system&#x0027;s DOF. This study paves the way for the application of OR-PAM technology in ophthalmic disease research.