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With a perfectly uniform illumination, the amount and concentration of fluorophores in any (biological) sample can be read directly from fluorescence micrographs. However, non-uniform illumination in optical micrographs is a common, yet avoidable artefact, often caused by the setup of the microscope, or by inherent properties caused by the nature of the sample. In this paper, we demonstrate simple matrix-based methods using the common computing environments MATLAB and Python to correctdoi:10.1364/oe.26.017279 pmid:30119541 fatcat:j5iwxwivdzee3gkkokapxms3ge