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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Proposals
The imaging parameters of the Imaging Setup Tool are defined for the dyes indicated in the panel:
Configure your experiment. Based upon this set of dyes used for staining the sample (see above)
Smart
Setup
computes four proposals with different goals:
−
Fastest
−
Best Signal
−
Smartest (Line)
−
Linear Unmixing
In the case of only a single type of fluorescent dye, all proposals may be identical.
The proposal
Fastest aims to record all dyes simultaneously as a single track with multiple channels
whenever possible. If the number of fluorescent dyes exceeds the number of available detectors, Smart
Setup switches to a speed-optimized multi-channel strategy with frame-wise switching between the
acquisition tracks. This proposal favors speed over cross talk.
The proposal for
Best Signal follows a strategy that maximizes the efficiency of signal recording and also
minimizes the level of cross talk. Multi-color acquisitions using this strategy are typically performed as
sequential acquisitions of each dye species with frame-wise switching between tracks.
Smartest (line)
combines the advantages of
Fastest
and
Best Signal
. This proposal aims to minimize
the number of acquisition tracks as well as cross talk. This is achieved by recording dyes with low spectral
overlap as a single acquisition track. In addition,
Smartest (Line) improves the acquisition speed also by
performing line-wise switching between tracks whenever applicable.
Unlike in the other modes, data recorded with
Linear Unmixing (only available if Show all is active) will
contain substantial amounts of fluorochrome cross talk. This data is meant to be processed (Linear
Unmixing) in order to separate individual fluorochromes.