Autofocusing optical-resolution photoacoustic microscopy

Optical-resolution photoacoustic microscopy (OR-PAM) is a promising noninvasive biomedical imaging technology capable of achieving micrometer or even sub-micrometer resolution. Its performance, however, is highly dependent on precise focusing. Traditional focusing methods are operator-dependent, low precise, and time-consuming, which greatly limits the practical applicability of OR-PAM, especially for users without sufficient training. In this study, we developed an autofocusing optical-resolution photoacoustic microscopy (AFOR-PAM) system by combining 3D motorized scan, unified data acquisition and motion control through a System-on-Chip (SoC) setup and an autofocusing algorithm based on a modified Brenner gradient function. The entire focusing process is fully automated and does not rely on the operator's experience. Phantom experiments verified the autofocusing capability of the AFOR-PAM system, requiring only 11 iterations and 1.4 s to complete a focusing process. In vivo imaging of the mice ears further demonstrated that AFOR-PAM can rapidly achieve more precise and reliable focusing than that with the conventional amplitude-based method. Moreover, the system exhibits strong robustness even under low signal-to-noise ratio (SNR) conditions. These advantages suggest that the proposed AFOR-PAM system has the potential to significantly broaden the practical applications of photoacoustic imaging in clinical settings.