Scanning Parameters

High resolution scans for deconvolution

• Choose appropriate voxel sizes (sampling distances) in X/Y and Z direction (scan more slices and less accumulation per slice, if possible (in most cases, accumulation=2 is enough)
• set accurate offset (low background, but no green pixels) and PMT setting (almost no blue pixels). Otherwise the correction by deconvolution may not be correct
• In case of an offset set too high, negative noise may be filtered out and calculation may be wrong caused by incorrect distribution of gray values
• Scan: include all visible structures plus non-sharp slices plus several additional seemingly signal-free slices, because they may contain low signals needed for correct deconvolution

Determining the ideal sampling intervals for high resolution 3D imaging

• use the Nyquist rate and PSF Calculator from SVI
• "For Confocal Microscope images sampling distances may be up to 1.7 times the Nyquist ones. When large pinholes are used, up to 2 times larger" (cited from the SVI WIKI page on Nyquist rate and PSF calculator

Another calculator

The Java applet is taken from http://www.pasteur.fr/recherche/unites/Pfid.

BITPLANE sampling calculator (from BITPLANE, Switzerland)

• Sampling interval (voxel size) table (calculated values a bit less restrictive)