Chapter 1 The Resolution Challenge in 3D Optical Microscopy

This chapter discusses the theoretical principles of 3D microscopy with the widespread realizations of 3D microscopy.Based on the paraxial diffraction equations, it has been shown that conventional microscopes, when dealing with 3D fluorescent samples, provide sets of 2D images. These images of the different transverse sections of the 3D object contain, in addition to the sharp image of the in focus section, the blurred images of the rest of the specimen. The paraxial formalism has been generalized in a very simple way to a non-paraxial context, showing that the equations that govern non-paraxial imaging are similar to those that govern paraxial imaging. The only difference is the integral that provides the 3D PSF. Therefore, all the background acquired in many decades of study concerning paraxial imaging can easily be transferred to the non-paraxial regime.