The lack of tools for sex identification and assessment of gonadal development are hindering our ability to study the reproductive dysfunction of Arapaima gigas in captivity. This study initially aimed to validate a non-surgical endoscopy procedure to identify sex in juveniles and assess stage of ovary development in female broodstock under field operational conditions. Cannulation, assisted through the description of the genital anatomy, made ovarian biopsy possible to describe oocyte development from primary growth to pre-ovulation, providing a first classification scheme for oogenesis in the species including description of the micropyle morphology using scanning electron microscopy. Cannulation was also successfully performed without endoscopic guidance, which allowed monitoring of ovarian development along the reproductive season together with profiling of plasma sex steroids (17β-estradiol [E₂] and 11-ketotestosterone [11-KT] in females and males, respectively). The monitoring of our study population showed females paired with males in earthen ponds sexually matured and reached oocyte maturation during the spawning season. However, since no spawning was recorded, eggs had either been resorbed or released and not fertilized by the male. Plasma E₂ levels remained high in females, as expected in an asynchronous species during the spawning season with multiple batches of oocytes being recruited. Plasma 11-KT showed a tendency to decrease, suggesting a male reproductive dysfunction or the end of the reproductive season with a lack of synchronization between sexes. In conclusion, endoscopy and cannulation are tools that can be promptly applied to aid sex identification, assessment of reproductive function, and overall broodstock management in wild and captive stocks. These tools will greatly help future studies looking at the effects of environmental, social, and hormonal cues on reproductive development with the aim of developing a spawning induction protocol for the species.