Method for determining the positions of a plurality of objects in a digital image by discriminating true positive positions of the plurality of objects from false positive candidate positions of the plurality of objects, the plurality of objects being configured to receive molecules comprising genetic information, the method comprising the following steps, preferably in the following order: a) determining a plurality of pixels in the digital image, each pixel having a local maximum intensity value with respect to the intensity values of a plurality of neighboring pixels of the respective pixel; b) determining the variance of the intensity value of each pixel having a local maximum intensity value with respect to the intensity value of the respective pixel having a local maximum intensity value and the intensity value of each of the plurality of the respective neighboring pixels; c) weighting the intensity value of each of the plurality of pixels having a local maximum intensity value by the respective determined variance of the intensity value; d) creating a histogram comprising the respective weighted intensity of each pixel of the plurality of pixels each having a local maximum intensity value; and e) determining a plurality of local minimums and the global maximum of the histogram; f) selecting one of the plurality of local minimums as a threshold value for discriminating true positive positions of the plurality of objects from false positive positions of the plurality of objects, wherein the one local minimum is selected under consideration of the position of the selected local minimum of the histogram relative to the position of the global maximum; and g) using the threshold value for discriminating the true positive positions of the plurality of objects from false-positive candidate positions of the plurality of objects. a) determining a plurality of pixels in the digital image, each pixel having a local maximum intensity value with respect to the intensity values of a plurality of neighboring pixels of the respective pixel; b) determining the variance of the intensity value of each pixel having a local maximum intensity value with respect to the intensity value of the respective pixel having a local maximum intensity value and the intensity value of each of the plurality of the respective neighboring pixels; c) weighting the intensity value of each of the plurality of pixels having a local maximum intensity value by the respective determined variance of the intensity value; d) creating a histogram comprising the respective weighted intensity of each pixel of the plurality of pixels each having a local maximum intensity value; and e) determining a plurality of local minimums and the global maximum of the histogram; f) selecting one of the plurality of local minimums as a threshold value for discriminating true positive positions of the plurality of objects from false positive positions of the plurality of objects, wherein the one local minimum is selected under consideration of the position of the selected local minimum of the histogram relative to the position of the global maximum; and g) using the threshold value for discriminating the true positive positions of the plurality of objects from false-positive candidate positions of the plurality of objects.