Photoacoustic microscopy in vivo using synthetic-aperture focusing technique combined with three-dimensional deconvolution.

Acoustic-resolution photoacoustic microscopy (ARPAM) plays an important role in studying the microcirculation system of biological tissues with deep penetration. High lateral resolution of ARPAM is achieved by using a high numerical aperture acoustic transducer. The deteriorated lateral resolution in the out-of-focus region can be alleviated by synthetic aperture focusing technique (SAFT). Previously, we reported a three-dimensional (3D) deconvolution ARPAM to improve both lateral and axial resolutions in the focus region. In this study, we present our extension of resolution enhancement to the out-of-focus region based on two-dimensional SAFT combined with the 3D deconvolution (SAFT+Deconv). In both the focus and out-of-focus regions, depth-independent lateral resolution provided by SAFT, together with inherently depth-independent axial resolution, ensures a depth-independent point spread function for 3D deconvolution algorithm. Imaging of 10 μm polymer beads shows that SAFT+Deconv ARPAM improves the -6 dB lateral resolutions from 65-700 μm to 20-29 μm, and the -6 dB axial resolutions from 35-42 μm to 12-19 μm in an extended depth of focus (DOF) of ∼2 mm. The signal-to-noise ratio is also increased by 6-30 dB. The resolution enhancement in three dimensions is validated by in vivo imaging of a mouse's dorsal subcutaneous microvasculature. Our results suggest that SAFT+Deconv ARPAM may allow fine spatial resolution with deep penetration and extended DOF for biomedical photoacoustic applications.