Method for background compensation of a digital image containing at least one first object, the at least one first object having received at least one molecule comprising genetic information, and at least one second object, the at least one second object not having received a molecule comprising genetic information, the at least one molecule being configured to receive a fluorescent compound, the digital image being determined by an optical imaging system during emission of electromagnetic radiation by the fluorescent compound, wherein the method comprises the following steps in the following order: a) determining first intensity values from the digital image for each object; b) determining second intensity values from the digital image for the at least one second object within a plurality of predetermined first areas, wherein the predetermined first areas comprise the at least one second object and one or more predetermined second areas that do not contain the at least one second object; c) computing median intensity values from the second intensity values for each of the plurality of predetermined first areas; c1) arranging the computed median intensity values in a topological grid structure based on a position of the at least one second object within the plurality of predetermined first areas, wherein the topological grid structure comprises at least one empty element at positions of the one or more predetermined second areas; c2) applying a region growing approach to the computed median intensity values arranged in the topological grid structure of the plurality of predetermined first areas to fill the at least one empty element in the topological grid structure; d) interpolating the median intensity values to the size of the original digital image; e) computing background compensated intensity values for each object by subtracting the interpolated median intensity values from the determined first intensity values. a) determining first intensity values from the digital image for each object; b) determining second intensity values from the digital image for the at least one second object within a plurality of predetermined first areas, wherein the predetermined first areas comprise the at least one second object and one or more predetermined second areas that do not contain the at least one second object; c) computing median intensity values from the second intensity values for each of the plurality of predetermined first areas; c1) arranging the computed median intensity values in a topological grid structure based on a position of the at least one second object within the plurality of predetermined first areas, wherein the topological grid structure comprises at least one empty element at positions of the one or more predetermined second areas; c2) applying a region growing approach to the computed median intensity values arranged in the topological grid structure of the plurality of predetermined first areas to fill the at least one empty element in the topological grid structure; d) interpolating the median intensity values to the size of the original digital image; e) computing background compensated intensity values for each object by subtracting the interpolated median intensity values from the determined first intensity values. <EOS>