Advancing photoacoustic microscopy via single-source triple-beam interference excitation

Photoacoustic microscopy (PAM) uniquely combines optical resolution with ultrasonic penetration capabilities. However, conventional focused Gaussian beam excitation presents inherent limitations in detection visibility and spatial resolution. Although increasing optical NA can partially mitigate these constraints, it inevitably compromises working distance (WD) and depth of field (DoF) and thus poses challenges for large-volume imaging. To address this fundamental multi-physics trade-off, we propose a triple-beam interference excitation strategy that enhances PAM performance while preserving extended WDs and DoFs. Through precise angular modulation of triple coherent excitation beams generated from a single source and optimizing their interference fringe patterns, the destructive interference of photoacoustic signals along the detection axis can be effectively disrupted. This wavefront source manipulation enables a sixfold enhancement in axial detection visibility compared to conventional Gaussian beam-based PAM. Additionally, the implementation of a five-step phase-shifting algorithm on the interference pattern yields substantial two- to threefold improvements (beyond diffraction limits) in lateral and axial dimensions. The proposed methodology is validated through comprehensive theoretical modeling and test imaging experiments on phantoms and biological specimens.