LIFA TIRF

LIFA TIRF

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  • The LIFA is a camera-based fast FLIM system that operates in the frequency domain and is compatible with Total Internal Reflection Fluorescence (TIRF). TIRF-FLIM enables selective lifetime imaging down to 100nm from the coverslip, allowing exciting applications including quantitative membrane trafficking studies using FLIM-FRET and Multi-Color FLIM.

    The LIFA is easy to install - within the hour - and very easy to operate. The attachment is compatible with Leica®, Nikon®, Olympus®, and Zeiss® fluorescence microscopes. The LIFA is in addition well suited for high-content screening applications. The LIFA system has been judged "easy and highly quantitative" for a.o. FLIM-FRET studies.

    Reference: In this paper by the Beatson Institute for Cancer Research (Glasgow, UK) the LIFA has been applied in TIRF mode.

    Supported TIRF modules

    Different TIRF modules have been succesfully evaluated in combination to the LIFA system including Leica, Nikon, and Olympus. The laser-TIRF unit is mounted to the epi-attachment of the microscope. The system's directly modulated laser diodes are connected to the laser-TIRF unit using fiber optics. The TRiCAM modulated ICCD camera is mounted to the camera port of the wide field fluorescence microscope.

    TIRF users can upgrade their existing laser-TIRF setup with a LIFA system. For excitation a modulated laser source is required. Lambert Instruments offers the Multi-LASER, which is available with a range of excitation wavelengths among which 405nm, 445nm, 488nm, 515nm and 639nm.

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  • The LIFA - TIRF combination has the following key features:

    • Fast TIRF-FLIM, up to two lifetime images per second
    • Lifetime sensitivity 0-300 ns
    • Lifetime accuracy 50 ps r.m.s.
    • High quantum efficiency image intensifier
    • Easy integration into specialized image analysis pipelines
    • Time-lapses
    • FRET efficiency mapping
    • Multi-frequency acquisition & analysis for separation of multiple lifetimes
    • Polar (Phasor) plot inspection and separation of multiple lifetimes
  • Lambert Instruments provides the following LIFA models, which cover a wide range in spectral sensitivity and in lifetime sensitivity. Each model is based on a specific high-resolution Gen II or Gen III image intensifier.

    Model Photocathode Modulation frequency Lifetime sensitivity
    LIFA GenII S20 S20 1-120MHz 0-300ns
    LIFA GenII S25 S25 1-120MHz 0-300ns
    LIFA GenIII GaAs GaAs 1-120MHz 0-300ns
    LIFA X GenII S20 S20 0-100kHz, 1MHz-120MHz 0ns-1ms
    LIFA X GenII S25 S25 0-100kHz, 1MHz-120MHz 0ns-1ms
    LIFA X GenIII GaAs GaAs 0-100kHz, 1MHz-120MHz 0ns-1ms
    LIFA P GenII S20 S20 0-100kHz 100ns-1ms
    LIFA P GenII S25 S25 0-100kHz 100ns-1ms
    LIFA P GenII GaAs GaAs 0-100kHz 100ns-1ms
    LIFA P GenIII GaAsP GaAsP 0-100kHz 100ns-1ms

    For each of the models the following graph shows the spectral sensivity versus wavelength.

    photocathodes flim

    Click here for more information on the operating principle of image intensifiers for lifetime imaging.


    0-100 kHz
  • Support for automated microscopes

    Nikon Ti Eclipse.

     

    Support for XYZ stages

  • The LIFA - TIRF combination has the following applications:

    • Molecular interactions
    • Protein conformation
    • Biosensors
    • Oxygen concentration imaging in cells and tissue
    • NADH / FAD fluorescence dynamics
    • Viscosity imaging
    • Membrane dynamics
    • Membrane trafficking