Abstract
Time-resolved photon counting methods have a finite bandwidth that restricts the acquisition speed of techniques like fluorescence lifetime imaging microscopy (FLIM). To enable faster imaging, computational methods can be employed to count photons when the output of a detector is directly digitized at a high sampling rate. Here, we present computational photon counting using a hybrid photodetector in conjunction with multithreshold peak detection to count instances where one or more photons arrive at the detector within the detector response time. This method can be used to distinguish up to five photon counts per digitized point, whereas previous demonstrations of computational photon counting on data acquired with photomultiplier tubes have only counted one photon at a time. We demonstrate in both freely moving C. elegans and a human breast cancer cell line undergoing apoptosis that this novel multithreshold peak detection method can accurately characterize the intensity and fluorescence lifetime of samples producing photon rates up to 223%, higher than previously demonstrated photon counting FLIM systems.
Original language | English (US) |
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Pages (from-to) | 2748-2755 |
Number of pages | 8 |
Journal | ACS Photonics |
Volume | 9 |
Issue number | 8 |
DOIs | |
State | Published - Aug 17 2022 |
Keywords
- FLIM
- fluorescence lifetime
- multiphoton microscopy
- photon counting
- single-photon detection
ASJC Scopus subject areas
- Electronic, Optical and Magnetic Materials
- Biotechnology
- Atomic and Molecular Physics, and Optics
- Electrical and Electronic Engineering