Zeiss LSM 880, Operating Manual

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CHAPTER 1 - SYSTEM OPERATION
ZEISS Left Tool Area and Hardware Control Tools LSM 880
222 000000-2071-464 10/2014 V_01
5.3.4.5 PCH
In the PCH (Photon Counting Histogram) panel you
can determine concentrations and the molecular
brightness of molecules.
•
Click on the PCH tab to open the PCH control
panel (Fig. 289).
You have the following options:
−
You can enter a background value, if you
know the background in the
Background
display box. Enter a zero, if you assume no
background. The background is no fit
parameter.
−
You can choose between 1, 2 or 3 components by activating the
Components 1
,
2
and
3
buttons,
respectively for each activated component you can enter a brightness value in Hz in the Specific
brightness
enter boxes. If you don’t know the brightness, enter 1.
−
You can enter
Instrumental parameters
values in the respective
first order
,
second order
and
third order enter boxes. These values have to be determined in independent calibration
experiments using a defined dye solution with a known brightness. The parameters correct for the
deviation of the confocal volume from a true Gaussian distribution. Normally, only a correction for
the first order parameter is necessary and recommended. Hence, in the calibration fit you will keep
the first order parameter free and the second and third parameters fixed to zero. You can enter the
determined first order number and fix it for later measurements.
Fig. 289 Model panel, PCH tab