New method for determining the depth of field of microscope systems.

This paper presents new formulas to determine the depth of field (DOF) of optical and digital microscope systems. Unlike the conventional DOF formula, the new methods consider the interplay of geometric and diffraction optics for infinite and finite optical microscopes and for corresponding digital microscope systems. It is shown that in addition to the well understood parameters such as numerical apertures, focal length, and light wavelength, system components such as aperture stops also affect the DOF. For the same objective lens, the DOF is inversely proportional to the size of the aperture stop, and it is proportional to the focal length of the ocular lens. It is also shown that under optimal viewing and operating conditions, the visual accommodation of human observers has no meaningful impact on DOF. The new formulas reported are useful for accurately calculating the DOF of microscopes.