An inspection system may generate first-step images of multiple sample regions after a first process step and generate second-step images of the sample regions after a second process step, where the second process step modifies the sample in at least one of the sample regions. The system may further identify one of the sample regions as a test region and at least some of the remaining sample regions as comparison regions, where the second-step image of the test region is a test image and the second-step images of the comparison regions are comparison images. The system may further generate a multi-step difference image by subtracting a combination of at least one of the second-step comparison images and at least two of the first-step images from the test image. The system may further identify defects in the test region associated with the second process step based on the multi-step difference image.