Serenity is a firefly that was formerly encased in a lump of amber that the Wayward Vagabond finds in the Skyship Base before it launches. Upon seeing the insect still alive and glowing in the amber, the Vagabond decides to use her to bring the light of democracy to Can Town. Later, the Vagabond releases her through use of the Appearifier and decides to name her something whimsical. The Vagabond actually has no idea if Serenity is a boy or a girl, or even if fireflies can be girls, but assumes the latter nonetheless.
She is intelligent enough to know that a countdown is rarely a good thing. She communicates via Morse code blinking her luminous rear end, and understands the passage of time. Her face seems to be stuck in the "blank stare" most of the time, which is used by many characters in MS Paint Adventures as well as the logo for the site.
A firefly that may have been Serenity appears in a lump of amber in the flash game at the start of Act 4.
In Cascade Serenity followed the fully prototyped Peregrine Mendicant to the trolls' Incipisphere with WV's almost dead body. In Act 6 Act 3, after many characters met up in a dream bubble and Dream Dirk flung Dream Roxy out of the bubble, Serenity departed with Dream Roxy. Together they arrived at post-scratch Derse where they were picked up by Dirk and taken back to post-scratch Earth via the Lotus Time Capsule.
Her namesake is the movie Serenity, which takes place on the titular spaceship after the final episode of the TV series Firefly. The ship Serenity is also known as a Firefly-class ship, so Serenity is quite literally a firefly.
Roxy renamed the firefly "doctor blinkbottom", and then "twinkly herbert", while also referring to it as being male, repeating an ongoing gag of animals being renamed and having their genders switched.
During Act 6 Act 5 Act 1, when Roxy falls asleep, Twinkly Herbert appears in her dream under the control of Calliope. Roxy is able to understand her Morse code messages while asleep.
==>: "Help! My friend is stuck inside the big can, inside a small house he built! You must hurry, he is not very bright and he doesn't understand it when I blink! Are you watching me? Oh no, don't blink me you don't understand blinking either!!! How do you people even exchange ideas without luminous rear ends!!!!!!"
Roxy naming Serenity "Twinkly Herbert" is a reference to an incomplete series of novels by Andrew Hussie named Wizardy Herbert. Not only does this compliment Roxy's love of wizards, but Zazzerpan and his Complacency originated as minor characters in the novel.