A method of determining a lateral misregistration between levels of a semiconductor structure comprises imaging at least one first alignment mark in a first level of the structure and at least one second alignment mark in a second level of the structure. A digital image of the first and second alignment marks is formed, each of which are defined by a set of points having an x-value and a y-value. The x-values and y-values of points defining the first alignment mark and points defining the second alignment mark are averaged to determine a center of the first alignment mark and a center of the second alignment mark. An x-coordinate and a y-coordinate of the center of the first alignment mark is subtracted from the respective x-coordinate and y-coordinate of the center of the second alignment mark to determine a lateral misregistration between the first level and the second level. Related methods of forming a semiconductor wafer, semiconductor assembles and metrology tools for use in implementing the methods are disclosed.