Multi-Wavelength FLIM

Multi-Wavelength FLIM

The principle of TCSPC FLIM can be extended to simultaneously detect in 16 wavelength channels. The optical spectrum of the fluorescence light is spread over an array of 16 detector channels. The TCSPC system determines the detection times, the channel numbers in the detector array, and the position, x, and y, of the laser spot for the individual photons. These pieces of information are used to build up a photon distribution over the time of the photons in the fluorescence decay, the wavelength, and the coordinates of the image. The principle of multi-wavelength FLIM is shown in the figure below.

Principle of multi-wavelength FLIM

As for single-wavelength FLIM, the result of the recording process is an array of pixels. However, the pixels now contain several decay curves for different wavelength. Each decay curve contains a large number of time channels; the time channels contain photon numbers for consecutive times after the excitation pulse. The result can also be interpreted as a number of FLIM images for different wavelength, as shown in the image below.

Multi-wavelength FLIM of a Convallaria sample. Images for 16 consecutive wavelength intervals, each 512×512 pixels, 256 time channels. Detected with MW-FLIM GaAsP detector, Aanalysis by SPCImage, Intensity-weighted lifetime of double-exponential decay model.

References for Multi-Wavelength FLIM

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  15. Rück, F. Dolp, C. Hülshoff, C. Hauser, C. Scalfi-Happ, FLIM and SLIM for molecular imaging in PDT, Proc. SPIE 5700 (2005)
  16. Rück, F. Dolp, C. Hülshoff, C. Hauser, C. Scalfi-Happ, Fluorescence lifetime imaging in PDT. An overview. Medical Laser Application 20, 125-129 (2005)
  17. Rück, Ch. Hülshoff, I. Kinzler, W. Becker, R. Steiner, SLIM: A New Method for Molecular Imaging. Micr. Res. Tech. 70, 403-409 (2007)
  18. Rück, C. Hauser, S. Mosch, S. Kalinina, Spectrally resolved fluorescence lifetime imaging to investigate cell metabolism in malignant and nonmalignant oral mucosa cells. J. Biomed. Opt. 19(9), 096005-1 to -9 (2014)

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