Many species exhibit variation in the color of their scales, feathers, or fur. Various forms of natural selection, such as mimicry, crypsis, and species recognition, as well as sexual selection, can influence the evolution of color. Eastern Indigo Snakes (Drymarchon couperi), a federally threatened species, have coloration on the sides of the head and the chin that can vary from black to red or cream. Despite significant conservations efforts for this species, little is known about its biology in the field. Past researchers have proposed that the color variation on the head and chin is associated with the sex of the individual. Alternatively, color might vary among individuals because it is controlled by genes that are under natural selection or neutral evolution. We tested these alternative hypotheses by examining whether coloration of the sublabial, submaxillary, and ventral scales of this species differed by sex or among clutches. We used color spectrometry to characterize important aspects of color in two ways: by examining overall color differences across the entire color spectrum and by comparing differences within the ultraviolet, yellow, and red colorbands. We found that Eastern Indigo Snakes do not exhibit sexual dichromatism, but their coloration does vary among clutches; therefore, the pattern of sexual selection leading to sexual dichromatism observed in many squamates does not appear to play a role in the evolution and maintenance of color variation in Eastern Indigo Snakes. We suggest that future studies should focus on determining whether color variation in these snakes is determined by maternal effects or genetic components and if color is influenced by natural selection or neutral evolutionary processes. Studying species that exhibit bright colors within lineages that are not known for such coloration will contribute greatly to our understanding of the evolutionary and ecological factors that drive these differences.