Abstract
Optical microscopy improves in resolution and signal-to-noise ratio by correcting for the system’s point spread function; a measure of how a point source is resolved, typically determined by imaging nanospheres. Optical-resolution optoacoustic (photoacoustic) microscopy could be similarly corrected, especially to account for the spatially-dependent signal distortions induced by the acoustic detection and the time-resolved and bi-polar nature of optoacoustic signals. Correction algorithms must therefore include the spatial dependence of signals’ origins and profiles in time, i.e. the four-dimensional total impulse response (TIR). However, such corrections have been so far impeded by a lack of efficient TIR-characterization methods. We introduce high-quality TIR determination based on spatially-distributed optoacoustic point sources (SOAPs), produced by scanning an optical focus on an axially-translatable 250 nm gold layer. Using a spatially-dependent TIR-correction improves the signal-to-noise ratio by >10 dB and the axial resolution by ~30%. This accomplishment displays a new performance paradigm for optoacoustic microscopy.
Highlights
Optical microscopy improves in resolution and signal-to-noise ratio by correcting for the system’s point spread function; a measure of how a point source is resolved, typically determined by imaging nanospheres
The optoacoustic point spread function (PSF) differs from the optical PSF in four ways: (1) optoacoustic signals exhibit a time profile carrying depth information, whereas optical signals are time-independent and represent a single spatial point; (2) the optoacoustic PSF is defined laterally by the optical focus and axially by the detector’s characteristics, while the optical PSF is determined only by the optical focus; (3) ultrasound detectors induce significantly stronger spatially-dependent aberrations to the recorded signals compared with optical detectors; (4) optoacoustic signals are bipolar, while optical signals have only positive intensities
The optoacoustic PSF constitutes in this regard the 3D spatial representation of signals from a point source incorporating (1) the optical impulse response describing the characteristics of optical excitation, (2) the spatial impulse response (SIR) capturing the spatiallydependent signal modification by the ultrasound detection, and (3) the spatially-invariant electric impulse response (EIR) embodying the signal digitization[28,29,30]
Summary
At positive defocus (Fig. 3f), frequencies showed a less pronounced pattern than at negative defocus, and high frequencies were lost as the lateral offset increased These TIR experiments with the GL detected higher frequencies of optoacoustic signals as well as a broader bandwidth for the transducer than the manufacturer’s pulse-echo characterization (see Supplementary Fig. 9). The joined and interpolated MAPs over the full 3D volume (Fig. 5g) as well as the side view of the central plane (Fig. 5h) showed an asymmetric sensitivity field along the central axis This convergence of simulation and experiment suggests that observed optoacoustic signal alterations and distortions can be attributed to internal reflections and interferences in the glass lens due to axial and lateral offsets of the signal’s origins. The TIR-SMF approach was again found to be superior to these alternatives in spatial representation of the data, noise reduction, and axial resolution improvement
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