Reproductive biology information is an important tool for fishery management actions such as the identification of spawning areas and the development of protective size limits, bag limits, and seasons. Such information for the management of Atlantic TripletailLobotes surinamensis is currently limited, particularly in the western Atlantic Ocean, as information regarding the reproductive biology of this species is sparse in the published literature. To this end, we determined the reproductive status of tripletail and compared the results of a nonlethal sampling method, plasma vitellogenin (VTG) analysis, with those of two traditional (lethal) methods, gonadosomatic index (GSI) and gonad histology. A total of 223 (122 male and 101 female) tripletail were sampled over 2 years near Jekyll Island, Georgia. Gonad histology indicated that 107 (94%) of the male tripletail were in the spawning-capable reproductive phase. Female tripletail were found in all reproductive phases, but only nine (8.9%) were in the spawning-capable phase. Plasma VTG was strongly related to GSI in females (R² = 0.832, n = 77), and female GSI differed significantly among reproductive phases (p < 0.0001). The estimated length at which 50% (L50) of female tripletail reached maturity was 463 mm; however, the L50 for male tripletail could not be determined because of the lack of immature fish within the study sample. Our study provides valuable information for the management of tripletail and indicates that a nonlethal approach (plasma VTG) may be useful for differentiating developing and spawning-capable females from males and from females in other reproductive phases.