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F(L)CS Data Acquisition
This makes it easier to allocate the images in LAS AF to the corresponding FCS data in
SymPhoTime.
18.4.5 FCS z-Stack
An FCS z-stack can be particularly useful for FCS measurements on membranes. To create
an FCS z-stack, follow these steps:
1. First acquire an xyz stack in the Setup Imaging step in the FCS Wizard ( Chapter 18.2 ) .
2. If necessary, optimize the measurement conditions in the
Setup FCS
step (
Chapter 18.3
).
3. Change to the Measurements step.
4. Use the image display to find the xy point in the z image stack at which you want to
generate the FCS z-stack.
5. In the z image stack, go to the top z plane that is of interest to you. Click on the desired
xy position and then on Add in Beam Path Settings ( Figure 179 ). Go to the plane below;
do not change the xy position, but click
Add
. Then go to the plane below that, etc.
Continue until you reach the lowest plane that is of interest to you.
If you wish, you can also acquire the z FCS stack from the bottom upwards.
6. Define the measurement duration, the number of repetitions at each point, and the
number of cycles as described in
Chapter 18.4.3
. The FCS instrument parameter settings
are identical at all measuring points.
7. Specify folder and file names as described in
Chapter 18.4.4
.
8. Click the Run FCS image acquisition button in LAS AF.
The maximum number of FCS measurements in a measurement series is 10,000.