Systems and methods for non-linear histogram segmentation for particle analysis are provided. In one embodiment, a method for analyzing particles comprises creating an initial two-dimensional histogram based on two selected parameters of the particles, filtering the initial two-dimensional histogram to generate a filtered two-dimensional image, detecting a plurality of seed populations in the filtered two-dimensional image, generating one or more linear contour lines, each having a plurality of contour points, to separate the detected seed populations, and adjusting the contour points in at least one of the linear contour lines to separate the detected seed populations.