Living Image 3.1

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H. 3D Reconstruction of Light Sources 240 Series or the IVIS Spectrum. For IVIS Spectrum or 200 Series data, if there is significant signal on the side of the subject, a larger angle of 70-85° can be used. Kappa Limits (DLIT) Kappa (κ) is a parameter that is searched during a reconstruction to determine the best fit to the image data. Small values of kappa tend to favor deeper sources, while large values favor more shallow sources. The limits on kappa are minimum of 0.1 and a maximum of 10. The default range for kappa is 0.5-4. Kappa is doubled at each iteration, so for a selected range of 0.5-4, the kappa values for each iteration would be 0.5, 1, 2. and 4. Choosing a large range for kappa produces the most reliable solution, but requires more analysis time. N Surface (FLIT) The number of surface intensity points to use in the reconstruction at a given source position. N Surface Limits (DLIT) This is the maximum number of surface intensity points to use in the reconstruction at a given wavelength. The range is 200 to 800 and the default is 200. The time required for reconstruction is shortest for smaller values of N (for example, 200). However, a large N value may give a more accurate result because more data are included in the fit. Starting Voxel Size (FLIT) Voxels are the small cubes of space inside a subject, each of which contains a quantity of fluorescent yield. The FLIT reconstruction begins with large voxels, specified by the starting voxel size (the length of a voxel cube side in mm). At each iteration, the algorithm reduces the size of the voxels by a factor of 2 until the optimum solution is determined. Starting Voxel Size Limits (DLIT) Voxels are the small cubes of space inside a subject, each of which contains a light source (much like a pixel in a 2D image). The DLIT reconstruction begins with large voxels, specified by the voxel size limit (the length of a side of the voxel cube in mm). At each iteration, the algorithm reduces the size of the voxel by a factor of two until the optimum solution is found. The voxel size limits are a minimum of five and a maximum of 10. The default range is set to 6-9 mm. A larger range of voxel limits ensures a more reliable solution, but requires more computational time. Voxel Size Increment (DLIT) This is the step increment in voxel size, stepping from the minimum voxel size limit to the maximum voxel size limit. For example, if the voxel size limit ranges from 6-9 mm, a voxel size increment = 1 gives four starting voxel sizes (6, 7, 8, and 9 mm). The default increment of 1 mm is usually adequate, however smaller increments can be used if you want to sample finer voxel sizes. Smaller increments will significantly increase the time required for reconstruction.
<strong>Living</strong> <strong>Image</strong> ® Software User’s Manual Background (FLIT) Choose this option to take the background fluorescence (for example, autofluorescence or non-specific probe in circulation) into account. Background fluorescence and fluorophore emission contribute to the photon density signal at the surface. The background fluorescence signal is modelled in order to isolate the signal due to the fluorophore only, where an average homogenous tissue background fluorescence yield is determined empirically. Background fluorescence contribution to the photon density at the surface is forwardmodelled. Simulated photon density data due to background at the surface is subtracted from the measured photon density so that the subsequent photon density used in the fit consists only of signal that is associated with the fluorophore. The background option specifies whether or not the background fluorescence should be fit. This is not necessary for the XFM-2, long wavelength data. It is important when there is non-specific dye in circulation or a lot of autofluorescence. Uniform Surface Sampling If this option is chosen, the surface data for each wavelength will be sampled spatially uniformly on the signal area. If this option is not chosen, the maximum ‘N surface elements’ will be sampled for the data. This means that the N brightest surface elements will be used as data in the reconstruction. Typically, non-uniform sampling is recommended if there is a single bright source, while uniform sampling is preferred if there are several scattered sources. NNLS Optimization + Simplex Optimization (DLIT) If NNLS Optimization + Simplex option is chosen, the software uses a linear programming algorithm to seed the solution, followed by the NNLS optimization. NNLS Weighted Fit Choose this option to weight the data in the NNLS optimization. 241
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Living Image 3.1

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