Continuity mistake: During "Brand New Day", at one point, Dr. Horrible stabs a picture of Captain Hammer attached to a dartboard with a handful of darts, all right in the center of the picture. A few shots later as Dr. Horrible sits, suddenly the darts are far-more spread out. (It's in the background and can be hard to see, but it's noticeable.)
Revealing mistake: During "Brand New Day", watch the background as Dr. Horrible appears as a giant and stomps down the city-street. As he takes his first step, a piece of concrete from building to his right falls off and hits the ground. However, if you look closely, it's obviously a quickly-made computer effect. As the piece of concrete hits the ground, it simply vanishes, and is replaced by a rather odd-looking animated "poof" of dust. The issue is the concrete piece was much bigger than the cloud of dust, so it's clear that the piece simply disappears right as the dust-poof appears.
Trivia: After the series became a major success upon release in 2008, director Joss Whedon teased that he would be doing a sort-of "sequel" at some point in the future. At one point, an outline and a number of songs for the sequel were written, though due to other obligations, Whedon had to push back production. As of 2018, the long-rumored sequel has yet to materialize, though everyone involved has continued to hint that a follow-up may still happen eventually.
Trivia: The film was written during the 2007-2008 Writer's Guild strike, which served as an inspiration for the production. Director/Co-Writer Joss Whedon, whom took part in the strike, wanted to make an independent project that he could create without having to deal with the red-tape of studios, unions, etc. Essentially to avoid the same sort-of issues that caused the strike in the first place.
Trivia: Was originally meant to be shot entirely with web-cams, with the characters reflecting on the events through dialog in video-blogs. However, after the story grew bigger during the writing process, it was ultimately decided that it would be filmed like a more traditional film or series, and the video-blogs became more of an occasional recurring motif than a main focus.