Adaptive fluorescence lifetime imaging with per-pixel signal optimization and flexible scanning

Area of Science:

• Biophotonics and advanced microscopy techniques.
• Development of novel imaging modalities for biological samples.

Background:

• Time-resolved fluorescence lifetime imaging microscopy (FLIM) offers enhanced biological imaging over steady-state methods.
• Modern FLIM faces challenges in achieving high resolution, signal-to-noise ratio (SNR), and speed in heterogeneous samples.

Purpose of the Study:

• To present fluorescence lifetime intensity-inverted imaging microscopy (FLI3M), an adaptive imaging technique.
• To demonstrate FLI3M's capability to balance imaging speed, signal strength, and sample integrity for uniform SNR imaging.

Main Methods:

• Developed FLI3M based on confocal laser scanning microscopy (CLSM).
• Implemented adaptive pixel dwell time adjustment using a priori intensity information from pre-scan.
• Supported flexible scanning patterns for dynamic imaging.

Main Results:

• Achieved uniform SNR imaging with up to eight-fold signal enhancement or reduced imaging time without SNR compromise.
• Demonstrated a 56% average improvement in fluorescence lifetime estimation reliability in low-SNR regions.
• Increased imaging speed by 27% to 53% across various biological samples.

Conclusions:

• FLI3M enables detailed resolution of optical fingerprints in complex biological environments.
• The adaptive FLIM system is a powerful tool for high-performance cellular imaging and FLIM-guided diagnostics.
• Established FLI3M for a wide range of biomedical applications.