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Related Experiment Video

Updated: Jun 3, 2026

Visualizing Protein Kinase A Activity In Head-fixed Behaving Mice Using In Vivo Two-photon Fluorescence Lifetime Imaging Microscopy

Visualizing Protein Kinase A Activity In Head-fixed Behaving Mice Using In Vivo Two-photon Fluorescence Lifetime Imaging Microscopy

Published on: June 7, 2019

Photon counting, censor corrections, and lifetime imaging for improved detection in two-photon microscopy.

Jonathan D Driscoll¹, Andy Y Shih, Satish Iyengar

¹Department of Physics, University of California at San Diego, La Jolla, CA 92093-0374, USA.

Journal of Neurophysiology

|April 8, 2011

Summary

We developed a fast photon counter for two-photon microscopy that improves signal-to-noise ratio by counting photocurrent pulses. This system can also be reconfigured for fluorescence lifetime imaging to differentiate fluorophores in vivo.

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Last Updated: Jun 3, 2026

Visualizing Protein Kinase A Activity In Head-fixed Behaving Mice Using In Vivo Two-photon Fluorescence Lifetime Imaging Microscopy

Visualizing Protein Kinase A Activity In Head-fixed Behaving Mice Using In Vivo Two-photon Fluorescence Lifetime Imaging Microscopy

Published on: June 7, 2019

In Vivo Two-Color 2-Photon Imaging of Genetically-Tagged Reporter Cells in the Skin

In Vivo Two-Color 2-Photon Imaging of Genetically-Tagged Reporter Cells in the Skin

Published on: July 11, 2019

Area of Science:

Biomedical Optics
Neuroimaging
Photon Counting

Background:

Two-photon microscopy offers high resolution for deep tissue imaging.
Traditional analog integration in photon detection can limit signal-to-noise ratio.
Accurate photon counting is crucial for advanced microscopy techniques.

Purpose of the Study:

To present a high-speed photon counter optimized for two-photon microscopy.
To enhance signal-to-noise ratio beyond analog methods.
To demonstrate the system's versatility for fluorescence lifetime imaging.

Main Methods:

Developed a high-speed photon counting system for photocurrent pulses.
Implemented an estimation algorithm to correct for dead time in photon detection.
Reconfigured the system in software for fluorescence lifetime imaging (FLIM).

Main Results:

The photon counting approach maximizes signal-to-noise ratio by avoiding analog integration limitations.
The dead time correction algorithm improves count accuracy.
Successfully distinguished spectrally similar fluorophores using FLIM in an in vivo microstroke model.

Conclusions:

The presented high-speed photon counter significantly enhances two-photon microscopy performance.
The system's adaptability to FLIM provides a powerful tool for biological research.
This technology aids in differentiating fluorophores for in vivo imaging applications.