Leica TCS SP8 SMD, User Manual

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FLIM Data Acquisition
Figure 142: Defining xy FLIM stacks
7. Define the duration of the individual FLIM images as described in Chapter 16.3.3 . If all
images are to have a comparable brightness, use the Acquire until max ___ photons/
pixel option. It enables corrections of intensity changes which are caused by the
emissions spectrum and by bleaching artefacts. If spectral intensity information is
required, use
Duration of each image or Repetitions .
8. Start the FLIM stack acquisition with the image acquisition button Run FLIM .
In LAS AF, you obtain a
λ
stack of averaged images and, in SymPhoTime, a series of FLIM
images.
16.3.8 Defining a Time Series of xy
λ
or xz
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FLIM Stacks
These modes are only available for systems that are also equipped with internal SP FLIM
detectors.
1. Select the specimen position.
2. In the Setup FLIM step on the Setup tab, select the Internal (SP FLIM) option
( Chapter 16.2.1 ).
3. In the Measurements step on the Acquisition tab, select either the Acquisition Mode
xy
λ
t or xz
λ
t as the scan mode. Optimize the instrument parameter setting for FLIM
measurement.
4. Define a file name in the Setup tab ( Chapter 16.3.2 ).
5. Open the Acquisition tab. If the correct SP FLIM detector is not yet selected, select it in
the PMT-Selection line. In Beam Path Settings , the detector selection cannot be
changed.
λ