CONFOCOR
3
ConfoCor 3
System Configuration
Carl Zeiss
02/2010 M60-1-0025
e
31
pupil filling will automatically be set to 100 %. The last entered value will be taken when switching back
to FCS mode.
•
It is recommended to set the pinhole diameter to a size corresponding to the used excitation
wavelength that corresponds to 1 AIRY unit:
−
458 nm 33 µm
−
488 nm 34 µm
−
514 nm 39 µm
−
561 nm 40 µm
−
633 nm 45 µm
Pinhole sizes are given in µm and AIRY units.
In the ConfoCor 3 you have one pinhole for both channels from the LSM.
Adjustment of Pinhole positions
The
Adjust Pinhole
panel is used to adjust the
beam path to the pinhole of the LSM. Adjustment
can be performed manually or automatically.
Manual adjustment is not recommended, since the
relevant procedure is very complex.
•
Open the
Adjust
Pinhole window (Fig. 36).
Press the tab again to close the window.
The pinhole is adjusted using a dye solution. The
general approach is to move the pinhole cyclically
in x and y until the intensity maximum is found. For
each excitation wavelength a suitable dye must be
used.
We recommend:
−
Rhodamine 6 Green (Rh6G) or Alexa 488 for 458, 488 and 514 nm,
−
Tetra-Methyl-Rhodamine (TMR) or Alexa 546 for 543 nm and
−
Cy 5 or Alexa 633 for 633 nm.
It is also recommended to work with a relatively concentrated solution (10
-6
mol/l) and low laser power to
achieve smooth intensity curves.
•
Activate the channel you want to use and that is appropriate for your settings by selecting it in the
check bullet. In one channel settings only the active used channel will be available. In two channel
settings both channels will be available.
Fig. 36
Pinhole adjustment panel
