Research fiber photometry system FLiP

neurosciencefiber opticfluorescence

Overview
The RWD Fluorescence Lifetime Photometry System (FLiP) combines Time-Correlated Single Photon Counting (TCSPC) with fiber photometry to monitor fluorescence lifetime changes in the brains of freely moving animals in real time. FLiP enables stable, quantitative in vivo measurements of neurotransmitter dynamics, protein–protein interactions and conformational changes by reporting fluorescence lifetime (τ), an intrinsic parameter largely independent of probe expression level, photobleaching and motion artifacts.

Why Fluorescence Lifetime?

• Lifetime is independent of fluorophore concentration and excitation power and reports local microenvironment and conformational changes.
• Provides quantitative, cross-sample comparable readouts without the need for baseline normalization.
• Suitable for stable long-term recordings in sleep studies, chronic disease models and longitudinal experiments.
• Enables sensitive detection of dynamic analyte changes using lifetime-based biosensors.

Two Mainstream Measurement Methods

• Time-Domain (TCSPC): Excites the sample with pulsed light and records temporal fluorescence decay. TCSPC timestamps individual photons to build decay histograms and extract lifetimes by exponential fitting. Offers high temporal resolution and superior SNR at low photon flux and can resolve multiple lifetime components with high precision.
• Frequency-Domain: Excites with sinusoidally modulated continuous light and derives lifetime from phase delay and modulation depth. Simpler to implement and fast, suitable for high photon flux samples or systems without pulsed lasers.

Comparison (Time-Domain vs Frequency-Domain)

• Lifetime measurement accuracy — Time-Domain (TCSPC): high; Frequency-Domain: good
• SNR for weakly fluorescent samples — Time-Domain (TCSPC): excellent; Frequency-Domain: relatively limited
• Resolution of multiple lifetime components — Time-Domain (TCSPC): high precision; Frequency-Domain: requires multi-frequency modulation or phasor analysis
• Acquisition speed — Time-Domain (TCSPC): relatively fast; Frequency-Domain: fast
• High photon flux samples — Time-Domain (TCSPC): good; Frequency-Domain: excellent
• Laser requirement — Time-Domain (TCSPC): pulsed laser; Frequency-Domain: modulated continuous light source

Highlights of the FLiP System

• System stability designed to minimize device-state fluctuations for reliable longitudinal comparisons.
• Low system noise to preserve signal integrity at weak fluorescence levels.
• Capability for long-term recordings with minimized baseline drift across consecutive weeks.
• Compatibility with a broad range of fluorescence lifetime biosensors.
• Direct quantitative analysis: planned standard-curve functionality to convert lifetime readouts toward concentration estimates.

Application Examples

• Neuromodulator measurements across animals and chronic time scales: lifetime readouts (e.g., GRAB ACh3.0) remain stable despite laser power and expression variability and can distinguish behavioral states.
• Mechanistic studies of psychiatric risk genes: FLiP applied to reveal abnormal striatal PKA dynamics in Akap11 mutant mice during freely moving recordings.
• Development and validation of novel lifetime sensors: e.g., LifeCamp calcium lifetime sensor enabling absolute measurement of transient and baseline calcium signals across cell types and behaviors.

Caractéristiques / spécifications techniques

• General system specifications
  • Excitation wavelength options: 488, 515, 560, 640, 670 nm (customizable)
  • Pulse frequency: 1–80 MHz
  • Lifetime measurement range: 1–10 ns
  • Sampling rate: 1–50 Hz
  • DIO count: 8
  • Voltage level: 3.3–5 V
  • Optical fiber core diameter: 200 or 400 µm
  • Numerical aperture (NA): 0.37
• Lifetime biosensor compatibility (examples with excitation wavelengths)
  • Ca2+: Tq–Ca–FLITS — excitation ~439 nm; tr-CCaMP — ~437 nm; tr-GCaMP — ~493 nm; tr-RCaMP — ~563 nm; RCaMP1h — ~560 nm; LifeCaMP — ~488 nm
  • ACh (acetylcholine): GRAB ACh3.0 — 488 nm; GRAB FLIM-ACh variants — 515 nm; GRAB WHalo-ACh-1.0 (dye-based) — 565 nm / 680 nm (JF dyes)
  • DA (dopamine): dLight3.8 — 488 nm; GRAB FLIM-DA1.0 — 515 nm
  • PKA: tAKARα, tAKARβ, tAKARγ — 488 nm; FLIM-AKAR — 488 nm; GRAB FLIM-PKA — 515 nm
  • PKC: CKAR3 — 488 nm
  • Extracellular glutamate: Rncp-iGluSnFR1 — 560 nm
  • Serotonin (5-HT): sDarken — 488 nm
  • Norepinephrine (NE): GRAB FLIM-NE1h / GRAB FLIM-NE1m — 515 nm; GRAB rNE3.0 — 560 nm; dye-based GRAB TMP-NE1.0 — ~690 nm; GRAB WHalo-NE — 565 nm / 680 nm (JF dyes)
  • 5-HT (alternate): GRAB FLIM-5-HT0.3 — 515 nm
  • CRF: GRAB rCRF1.0 — 570 nm
  • PACAP: GRAB rPACAP1.0 — 570 nm
  • Glucose: iGlucoSnFR-TS — 400 nm
  • Lactate: LiLac — 425 nm
  • ATP: qMaLioffG — 488 nm