A cell image segmentation method includes receiving a cell image, performing a nuclei initialization step to find an internal marker and an external marker to obtain a potential nuclei and a potential cell boundary, calculating a gradient map of the received cell image, performing a filtering step on the gradient map to generate a filtered gradient map, performing a nuclei detection step to obtain a segmented nuclei, and performing a nuclei validation step to obtain a valid nuclei. The nuclei initialization step includes performing a blob detection step to obtain a nuclei candidate, an outlier removal step to obtain the internal marker, a distance transform step to obtain a distance map, and a cell boundary initialization step to obtain the external marker. In another embodiment, a nuclear-to-cytoplasmic ratio evaluation method using the above cell image segmentation method is proposed.