Living Image 3.1

Living Image ® Software User’s Manual
8
Spectral Unmixing
Spectral Unmixing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 114
Spectral Unmixing Results Window . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 117
Spectral Unmixing Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Spectral Unmixing Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
The Living Image ® software uses spectral unmixing to distinguish the spectral
signatures of different fluorescent or bioluminescent reporters and calculate the
respective contribution of each on every pixel of an image. Use spectral unmixing to:
• Extract the signal of one or more fluorophores from the tissue autofluorescence.
Images can be acquired using epi-illumination (excitation light above the stage) or
transillumination (excitation light below the stage)
• Analyze bioluminescence images when more than one reporter is used in the same
animal model
Use the imaging wizard to setup the image sequence required for spectral unmixing.
(For more details on the imaging wizard, see page 23.) If you do not use the imaging
wizard to set up the image sequence, it is recommended that the image sequence include
images acquired using several filters that sample the emission or excitation spectra at
multiple points across the entire range. Make sure that the band gap between the
excitation and emission filters is sufficiently large (for example, >40 nm) so that the
excitation light does not leak through the emission filter where it can be detected by the
CCD.
113