Stiles added to her list of credits with films such as The Business of Strangers (2001), Mona Lisa Smile (2003), and The Omen (2006), and became known to audiences worldwide with her portrayal of Nicky Parsons in the Bourne franchise (2002–2016).

Her other notable film credits include Hamlet, State and Main (both 2000), O (2001), A Guy Thing (2002), Carolina (2003), The Prince & Me (2004), Edmond, A Little Trip to Heaven (both 2005), The Cry of the Owl (2009), Silver Linings Playbook (2012), Out of the Dark (2014), Blackway (2015), 11:55 (2016), Hustlers (2019) and Orphan: First Kill (2022).

[8] She also had small roles as Harrison Ford's character's daughter in Alan J. Pakula's The Devil's Own (1997) and in M. Night Shyamalan's Wide Awake (1998).

[9] Stiles was cast at the age of 17[6] for the role of Kat Stratford, opposite Heath Ledger in Gil Junger's 10 Things I Hate About You, an adaptation of The Taming of the Shrew set in a high school in Seattle.

Her next starring role was in Down to You (2000), which was panned by critics, but earned both her and her co-star Freddie Prinze, Jr. a Teen Choice Award nomination for their on-screen chemistry.

The second was in the Desdemona role, opposite Mekhi Phifer, in Tim Blake Nelson's O (2001), a version of Othello set at a boarding school.

Stiles's next commercial success was in Save the Last Dance (2001) as an aspiring ballerina forced to leave her small town in downstate Illinois to live with her struggling musician father in Chicago after her mother dies in a car accident.

Stiles also appeared opposite Stockard Channing in the dark art house film The Business of Strangers (2001) as a conniving, amoral secretary who exacts revenge on her boss.

Between the Bourne films, she appeared in Mona Lisa Smile (2003) as Joan, a student at Wellesley College in 1953, whose art professor (Julia Roberts) encourages her to pursue a career in law rather than become a wife and mother.

Critic Stephen Holden called her one of cinema's "brightest young stars",[12] but the film met with generally unfavorable reviews.

Critic Dennis Harvey wrote that Stiles was "wasted"[14] and Holden called her "a serious actress from whom comedy does not seem to flow naturally".

[22] While Stiles performed in a school play in fourth grade, Bob McGrath of Ridge Theater in Manhattan, a friend of her parents, needed an actor for a nonspeaking role.

In the summer of 2002, she performed on stage in Eve Ensler's The Vagina Monologues,[23] and appeared as Viola, the lead role in Shakespeare in the Park's production of Twelfth Night with Jimmy Smits.

[24] In 2004, she made her West End stage debut opposite Aaron Eckhart in a revival of David Mamet's play Oleanna at the Garrick Theatre.

[25][26] She reprised the role of Carol in a 2009 production of Oleanna, directed by Doug Hughes and co-starring Bill Pullman at the Mark Taper Forum.

[28] Stiles was to play Jeannie in a production of Neil LaBute's Fat Pig directed by the playwright beginning in spring 2011,[29] but the show was postponed indefinitely.

She works at an office and also as a call girl to make ends meet on an otherwise meager income fighting to protect her son from the collision between her complicated past and tenuous present.

[42] Stiles made her writing and directorial debut with Elle magazine's short Raving starring Zooey Deschanel.

In January 2004, Marie Claire featured Stiles's trip to witness conditions at the Berks County Youth Center in Leesport, Pennsylvania.