Introduction

Time-Correlated Single Photon Counting (TCSPC) is the “gold-standard” method for measuring fluorescence lifetime but is unsuitable for bright ambient light conditions, thus making it impractical in clinical applications. The invention described here ensures that the fluorescence and illumination light photons are temporally separated and the fluorescence signal is free from background light contamination.

Technical features

A pulsed bright light source is used to provide periodic illumination of the field of view. The TCSPC acquisition is out of phase with the bright light illumination, i.e. the measurement starts when the light source is turned off and is stopped before a new on-off cycle. This method guarantees that fluorescence and background photons are temporally resolved and the fluorescence signal is free from bright background light, which makes TCSPC measurements possible and practical under bright background conditions.
The impact and applicability of this method in clinical procedures is further increased by using a fibre-optic probe for excitation light delivery and fluorescence collection. Fluorescence lifetime maps can be created and displayed from single point fibre-based measurements by superimposing a visible guiding beam with the fluorescence excitation beam, which creates a visible reference onto the sample that can be imaged and segmented in real time.
The method has been validated in laboratory on biological samples (TRL4).
At the moment we are completing the ethical procedures required to test the technology in a clinical environment.

Possible Applications

• Clinical diagnostics;
• Surgical guidance;
• Therapy follow-up.

Advantages

• TCSPC measurements in the presence of a bright background;
• TCSPC measurements in clinical settings;
• Real-time acquisition and processing;
• Imaging with a single optical fiber;
• Fluorescence lifetime displayed as augmented reality.