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Cascaded spatial frequency modulation imaging architectures for enhanced resolution multiphoton imaging in photon counting regimes
Scarbrough, Daniel W.
Scarbrough, Daniel W.
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2023
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2025-06-24
Abstract
SPatIal Frequency modulation Imaging (SPIFI) is a well-established structured illumination imaging technique that has demonstrated enhanced resolution imaging with an extended illumination source and single-pixel detection. This thesis presents work done to improve modulation scan times as well as enhance performance in multiphoton microscopy.
The standard implementation of SPIFI uses a spinning disk to impart the spatial frequency modulation onto a line cursor. This architecture is simple to implement in a laser microscope, but is limited in speed and stability due to using a DC motor to spin a patterned disk. As an alternative, the use of polygonal scan mirrors was investigated. These mirrors operate at high speed and stability, but required development of a new SPIFI modulation architecture. The resulting systems showed a reduction in scan times of approximately two orders of magnitude while retaining enhanced resolution capability.
For multiphoton modalities which require ultrafast laser pulses, a new method of pulse characterization with SPIFI was investigated. While not completely successful, it led to a method for rapid dispersion compensation optimization as well as a collaboration with promising initial characterization results. The dispersion optimization system coupled with a spinning disk SPIFI system was used to optimize for dispersion through the full microscope system, demonstrating enhanced resolution imaging with pulse energies on the order of nanojoules without significant processing.
To pursue further resolution enhancement, photon counting SPIFI was demonstrated using a custom software application to provide a GUI interface and rapid, multi-threaded processing. To achieve stable modulation signals and rapid acquisition, the polygonal scan mirror architecture was used for this multiphoton microscope. Signals were acquired and processed for photon counting at a rate of 500 Hz with a simple USB oscilloscope. With this system, fourth order SPIFI enhanced images were acquired with 1.75 seconds of captured exposure per line and nanojoule pulse energies.
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