We present the first record of post-release follow-up and monitoring of a rehabilitated two-toed sloth (Choloepus hoffmanni), first record of freezing behavior and first antagonistic interaction of the genus Choloepus with an owl (Pulsatrix perspicillata). Two-toed sloths are nocturnal and arboreal mammals which survival relies on their capability to remain undetected by predators. Unfortunately, in the Guayas province of Ecuador, they are the among the species most abundantly received in rehabilitation centers. The liberation of animals back to the wilderness is the main goal of rehabilitation, and the follow-up, or, post-release human presence/support of animals, facilitates their reestablishment in their natural habitat. Follow-up, direct observation and Bluetooth-based monitoring of the sloth secured the survival of the two-toed sloth. The range of detectability of the device used indicates its suitability for tracking low-mobility animals. After the first five days, the number of trees used per day increased, and 21 trees within 1152 m² were visited. Daylight and movement time range showed correlation towards detectability. The follow-up effort permitted to maintain safe the two-toed sloth during 12 days after release. Due to the difficulty monitoring nocturnal animals, economic constraints in conservation, accessibility, and safety of the animals, biodegradable Bluetooth based backpacks are recommended to ease the location of the animal and support its survival in the wild.