Abstract: A method for overlay monitoring including: obtaining a secondary electron image and a backscattered electron image of as area of the substrate in which an array of first structural elements are positioned at a surface of the substrate and a second array of second structural elements are positioned below the first array; determining locations of the first structural elements within the secondary electron image; defining regions of interest in the backscattered electron image, based on the locations of the first structural elements; processing pixels of the backscattered electron image that are located within the regions of interest to provide a backscattered electron representation of a second structural element; and calculating an overlay error based on location information regarding the second structural element within the backscattered electron representation of the second structural element and on location information regarding of at least one first structural element in the secondary electron image.