Excitation-resolved imaging

Excitation-resolved imaging Tunable continuum source... 

Conventional TCSPC equipment has been successfully employed in LSM for fluorescence spectroscopy on discrete microscopic volumes [18, 19] and for fluorescence lifetime imaging at a low acquisition speed [1], The use of conventional TCSPC equipment for imaging results in very long acquisition times, several to many minutes per (time-resolved) image. Importantly, operating the TCSPC detection system at too high detection rates, above 5% of the excitation frequency, results in distortion of the recorded decay curve [20],... 

Fig. 7 Set-up for time-resolved imaging of 96-microwell plate formats, a Fast gateable CCD-camera, b Optical emission filter, c Light-guiding adapter, consisting of 96 optical fibers, d 96-microwell plate (black with transparent bottom), e Optical excitation filter, f Pulseable LED array with 96 light emitting diodes...

The combination of different fluorescent metal indicators with inert luminescent reference beads consisting of poly(acrylonitrile) containing Ru(dpp)3 leads to a sensor array in a microwell plate format, suited for ratiomet-ric time-resolved imaging [95]. The data can be acquired with the help of the f-DLR method (for details see Sect. 2.3). A cross-reactive sensor array was arranged for the determination of mixtures of calcium(II), copper(II), nickel(II), cadmium(II), and zinc(II) ions by nine different commercially available fluorescent indicators (Table 3). For a successful application, it is mandatory that all luminophores can be excited at the same wavelength range between 400 and 500 nm, and that the excitation and emission spectra of all indicators overlap with those of the reference dye encapsulated in the nanobeads. 

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