We propose a straightforward implementation of two-photon image scanning microscopy (2PE-ISM) that, by leveraging our recently introduced single-photon avalanche diode (SPAD) array detector and a novel blind image reconstruction algorithm is shown to dramatically improve the optical resolution of two-photon imaging, in various test samples. We show how our computational ISM approach is able to adapt to changing imaging conditions, thus ensuring optimal image quality. We also show how our recently introduced blind deconvolution approaches can be integrated into the image reconstruction workflow to further improve the image quality.
Photo-Acoustic Microscopy (PAM) has raised high interest in in-vivo imaging due to its ability to preserve the near-diffraction limited spatial resolution of optical microscopes, whilst extending the penetration depth to the mm-range. Another advantage of PAM is that it is a label-free technique – any substance that absorbs PAM excitation laser light can be viewed. However, not all sample structures desired to be observed absorb sufficiently to provide contrast for imaging. This work describes a novel imaging method that makes it possible to visualize optically transparent samples that lack intrinsic photo-acoustic contrast, without the addition of contrast agents. A thin, strongly light absorbing layer next to sample is used to generate a strong ultrasonic signal. This signal, when recorded from opposite side, contains ultrasonic transmission information of the sample and thus the method can be used to obtain an ultrasound transmission image on any PAM.
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