Image of a resolution test target that was tilted by ∼1.5° on the microscope stage. The spacing of the gratings in micrometers is marked on the specimen. Because of the tilt, the defocus varies across the image as indicated by the orange scale. The specimen-to-lens distance increases from right to left. Note that the black–white bars of the 0.29-µm grating are fairly distinct at the left end but smeared to an average gray on the right with a 60× (NA = 1.4) objective, 546-nm illumination.