Decreasing SNR is shown using a cell with 250 intensity units (iu) and no background (0 iu) in three scenarios of increasing s.d. (in iu) of background Gaussian noise: 0 (a), 50 (b) and 200 (c). The effect of decreased CR is displayed using a simulated cell in high background (200 iu) with increasing noise s.d.: 0 (d), 50 (e) and 200 (f). The effect is shown for three increasing noise levels: 0 noise (a versus d), 50 noise s.d. (b versus e) and 200 noise s.d. (

Intra-cellular signal heterogeneity that can lead to cell over-segmentation when the same cell yields several detections is simulated by a cell with nonuniform distribution of the labeling marker or nonlabel retaining structures (g). Signal texture can also be linked to the process of image formation, in this case shown using a simulated cell image imaged by PhC microscopy (h). (i) Signal heterogeneity between cells, shown by simulated cells with different average intensities can be a result of, for instance, different levels of protein transfection, non-uniform label uptake, or cell cycle stage or chromatin condensation, when using chromatin-labeling techniques.

Spatial resolution that can compromise the accurate detection of cell boundaries is displayed using a cell captured with increasing pixel size, i.e., with decreasing spatial resolution: full resolution (j), half resolution (k) and one fourth of the original full resolution (l). (m,n) Irregular shape that can cause over/under-segmentation, especially when the segmentation methods assume simpler, non-touching objects, is displayed using a simulated cell with highly irregular shape under two background noise s.d. situations: 0 (m) and 100 (n).T