A key has been developed for identifying the pupal exuviae of 132 taxa of chironomid midges collected in Everglades National Park, as well as 18 additional species from freshwater habitats adjacent to the Park. Descriptions and illustrations are based upon voucher specimens from extensive collections of chironomid pupal exuviae for faunal surveys and biomonitoring research conducted in ENP and surrounding freshwater areas from 1998 to 2007. The key includes taxonomic comments for confirming identifications, as well as brief summaries of the distribution and ecology of each species in southern Florida waters. Information is also provided on the morphology of chironomid pupal exuviae, recommended references for identifying pupal exuviae, techniques for making slides, and methods to confirm proper identification.