I am not getting much compression of the z-blur with a measured PSF, why?

• There is a problem with the measured PSF:
  • bead images were saturated or undersampled.
  • beads have formed aggregates.
  • beads were moving while being imaged.
  • insufficient signal from beads causing inaccuracies in the averaging procedure.
  • strongly varying background.
• The conditions under which the PSF was measured do not match the imaging conditions.
  • The most important parameter is the medium refractive index. To exclude magnification calibration problems, best record the bead images at the same sampling density as the specimen. To check whether there is a matching problem between the measured PSF and the specimen data, deconvolve the specimen with a theoretical PSF. If the result is better then there can be a matching problem.