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After beginning his career in sitcoms, Whedon wrote the poorly received horror comedy film Buffy the Vampire Slayer (1992)—which he later adapted into the acclaimed television series of the same name—co-wrote the Pixar animated film Toy Story (1995), and wrote the science fiction horror film Alien Resurrection (1997). After achieving success as a television showrunner, Whedon returned to film to write and direct the Firefly film continuation Serenity (2005), co-write and produce the horror comedy film The Cabin in the Woods (2012), and write and direct the Shakespeare adaptation Much Ado About Nothing (2012). For the Marvel Cinematic Universe, Whedon wrote and directed the ensemble superhero film The Avengers (2012) and its sequel Avengers: Age of Ultron (2015). He also co-wrote the DC Extended Universe superhero film Justice League (2017), for which he also served as director for reshoots, replacing Zack Snyder (who retained directorial credit).

Whedon has also worked as a composer (notably for the Buffy the Vampire Slayer musical episode "Once More, with Feeling" and Dr. Horrible's Sing-Along Blog) and comic book writer, both for comic book continuations of television series he created and for established franchises, such as Astonishing X-Men.

Beginning in July 2020, multiple actors, producers, and writers spoke out against Whedon following the toxic workplace environment he had allegedly created in many of his projects. Whedon has denied any wrongdoing, while acknowledging that he can be "confrontational."