3:45 PM - 4:00 PM
[21p-S422-8] Enhanced Speed in Frequency Division Multiplexed Fluorescence Confocal Microscopy
Keywords:fluorescence confocal microscopy,frequency division multiplexing,high-speed imaging
Fluorescence imaging using radiofrequency-tagged emission (FIRE) is an emerging technique that enables much higher imaging speed (namely, temporal resolution) in fluorescence microscopy compared to conventional fluorescence imaging techniques such as confocal microscopy and image-sensor-based wide-field microscopy. It works based on the principle that it uses multiple intensity-modulated fields in an interferometric setup as excitation fields and applies frequency-division multiplexing to fluorescence signals. Unfortunately, despite its high potential, FIRE has limited imaging speed due to signal bandwidth limited by that of an acousto-optic deflector (AOD) employed in the setup, which is typically 100-200 MHz for the spectral range of fluorescence excitation (400-600 nm). Here we present a method named dual-AOD FIRE to overcome the limitation and thus to achieve higher imaging speed than the prior version of FIRE. Our method achieves an increase in signal bandwidth by a factor of two, which enables the imaging speed limited by the lifetime of the target fluorophore rather than the imaging system itself. Due to its unprecedentedly high-speed performance, our method has a wide variety of applications in cancer detection, drug discovery, and regenerative medicine.
We found that our dual-AOD FIRE suffers from image artifact due to inter-pixel crosstalk when the number of pixels in parallelized direction is increased. We will demonstrate a method to cancel out such a crosstalk by appropriately controlling the driving signals to the AODs and high-speed fluorescence imaging at a frame rate of 16,000 frames/second by the dual-AOD FIRE.
We found that our dual-AOD FIRE suffers from image artifact due to inter-pixel crosstalk when the number of pixels in parallelized direction is increased. We will demonstrate a method to cancel out such a crosstalk by appropriately controlling the driving signals to the AODs and high-speed fluorescence imaging at a frame rate of 16,000 frames/second by the dual-AOD FIRE.