Abstract: An array of localized auto-focus sensors provides direct measurement of the working distance between each microscope column in the array and the substrate being imaged below. The auto-focus sensors measure the working distance between each column and the imaging substrate as it passes over a point on the substrate to be imaged. The working distance measurement from the sensors is input into a control system, which in turn outputs the required working distance adjustment to the microscope column. The control system independently adjusts microscope working distance and/or physical distance of an individual microscope column in a multi-column microscope based on auto-focus sensor input. The individual microscope columns in the multi-column microscope can also be used as the auto-focus sensor itself.