Point-Spread Function retrieval for fluorescence microscopy

In this paper we propose a method for retrieving the Point Spread Function (PSF) of an imaging system given the observed image sections of a fluorescent microsphere. Theoretically calculated PSFs often lack the experimental or microscope specific signatures while empirically obtained data are either over sized or (and) too noisy. The effect of noise and the influence of the microsphere size can be mitigated from the experimental data by using a Maximum Likelihood Expectation Maximization (MLEM) algorithm. The true experimental parameters can then be estimated by fitting the result to a model based on the scalar diffraction theory with lower order Spherical Aberration (SA). The algorithm was tested on some simulated data and the results obtained validate the usefulness of the approach for retrieving the PSF from measured data.