Other methods of segmentation and background estimation. (a) Frame difference segmentation: The segmentation of foreground cells based on frame differences. It can be observed that frame-difference segmentation does not yield a satisfactory segmentation result. Segmented cells end up as scattered pixels without any recognizable shape. (b) Morphological background estimation/subtraction: Estimated background and segmentation using morphological operators. Cells are not meaningfully segmented in the foreground that is obtained by applying morphological background estimation/subtraction of . Scale bar is 20 μm.