Zeiss LSM 880, Operating Manual

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CHAPTER 1 - SYSTEM OPERATION
LSM 880 Left Tool Area and Hardware Control Tools ZEISS
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5.2.15.4 Optimize Sectioning and Step
Expand the Optimize Sectioning and Step field
by pressing the triangle (Fig. 196). The
Optimize
Sectioning and Step function helps to set the
optimal
Interval
. It is in this respect redundant to
the Optimal button of the First/Last and Center
modes. If more than one channel is used, the
channel detecting the shortest spectral range
determines the optimal interval.
The percentage of overlap of subsequent image
slices is indicated for each channel and each track.
A 50% overlap is considered to be optimal
representing a 2fold Nyquist setting.
The function
Match Pinhole (Fig. 197) sets the
pinhole to keep this optimal interval and in
addition have the slice thickness for all detection
channels approximately the same. This typically
results in slightly thicker slices for channels
detecting the longer wavelength range. In case the
channels are assigned to different tracks and a
Frame wise Multitracking scheme is applied, the
pinhole diameter is set for each track such that the
values of the resulting optical sections from the
different channels are identical and have double
the value of the optimal interval.
Note that the settings of the pinhole
might not result in matching optical
sections. This is then due to the chosen
objective (very low NA) which would
require a smaller setting of the pinhole
that is mechanically not possible. In this
case chose a higher magnifying objective
or start with a larger pinhole setting for
the channel detecting the shortest
wavelength range.
Pressing the Undo button (only available in the SIM and LSM imaging modes) resets to the previous
value.
Pressing the
X:Y:Z = 1 button matches the settings in Z to the settings in X and Y. This produces a
cubical voxel.
Fig. 196 Optimize sectioning and Step –
appearance for the LSM imaging
mode
Fig. 197 Optimize sectioning and Step –
appearance for the LSM imaging
mode pinholes matched to
generate equal slice thickness