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Troubleshooting
• ATTO655 for 640 nm
• Rhodamine 6G for 532 nm or fluorescein (in a solution with a high pH value)
• ATTO488 for 470-nm excitation
• The molecular brightness is the count rate of the detectors divided by the number of
molecules present in the detection volume. It is displayed in the FCS preview. In general,
a 10-nM solution is used. For a day-to-day comparison, use the same laser power, ideally
controlled using a power meter. A more detailed description of FCS measurements is
provided in the point measurement section.
31.2.1
Causes for Decreased Performance
If a performance drop is noticed, the origin of the performance drop needs to be determined
first. A drop in system performance may be caused by:
• Decreasing laser power:
Check the laser intensity with a laser power meter. If the laser power from PQ lasers is
too low, either the laser coupling unit has to be readjusted or the laser needs to be
repaired or replaced. In this case, please contact customer support.
•
Decreasing detection sensitivity:
• Misalignment of the TCS SP8: Contact Leica Service.
• Misalignment of the fiber decoupler for the fluorescent light: Contact Leica Service.
• Misalignment of the detector(s): In this case, please contact Leica customer support.
31.3
No FLIM Image is Displayed During Measurement
Can laser light be seen over the objective during measurement?
If not:
• Is the shutter open in LAS AF?
• Has the laser been activated by moving the corresponding slider?
• Is the multifunction port in the correct position?
• Is the pulse repetition rate of the pulsed lasers correct? This can be checked in the Time
Trace preview window. When starting a measurement, at least background noise should
be displayed (see Figure 148 at the right in Chapter 17.8). If the repetition rate is equal to
0, check whether the SYNCHRONIZATION is set to external on the PDL800B or PDL800D
and whether the laser emission lamp is illuminated during scanning.
If yes:
• Do you see a dark count rate when activating the Time Trace tab in the Preview
window? If not, check to ensure that the detectors are still activated. The DSN 102 (if
present) deactivates a detector automatically if the count rate exceeds a certain limit. In
250
Observe additional manuals
For more information, refer to the manual for the PDL800B or D.
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