  Light microscopy as well as image acquisition and processing suffer from
physical and technical prejudices which preclude a correct interpretation of
biological observations which can be reflected in, e.g., medical and
pharmacological praxis. Using the examples of a diffracting microbead and
fluorescently labelled tissue, this article clarifies some ignored aspects of
image build-up in the light microscope and introduce algorithms for maximal
extraction of information from the 3D microscopic experiments. We provided a
correct set-up of the microscope and we sought a voxel (3D pixel) called an
electromagnetic centroid which localizes the information about the object. In
diffraction imaging and light emission, this voxel shows a minimal intensity
change in two consecutive optical cuts. This approach further enabled us to
identify z-stack of a DAPI-stained tissue section where at least one object of
a relevant fluorescent marker was in focus. The spatial corrections (overlaps)
of the DAPI-labelled region with in-focus autofluorescent regions then enabled
us to co-localize these three regions in the optimal way when considering
physical laws and information theory. We demonstrate that superresolution down
to the Nobelish level can be obtained from commonplace widefield bright-field
and fluorescence microscopy and bring new perspectives on co-localization in
fluorescent microscopy.
