The success of meiosis depends on intricate coordination of a series of unique cellular processes to ensure proper chromosome segregation. Many proteins involved in these cellular events are directly or indirectly associated with chromosomes, especially those required for homologous recombination. These meiotic processes have been explored extensively by conventional light microscopy. However, many features of interest, such as chromatin organization, recombination nodules, or the synaptonemal complex are beyond the resolution of conventional wide-field microscopy. Moreover, in most sample preparation techniques for light microscopy, meiotic cells are squashed, which destroys the spatial organization of the nucleus. Here, I describe a protocol to analyze maize meiotic chromosomes by three-dimensional structured illumination microscopy (3D-SIM), a recently developed high-resolution microscopy technique. This protocol can be used to examine protein localizations at a high resolution level by immunofluorescence.