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Biel began her career as a vocalist appearing in musical productions until she was cast as Mary Camden in the family drama series 7th Heaven (1996–2006), in which she achieved recognition.

In 1997, Biel won the Young Artist Award for her role in the drama film Ulee's Gold. She received further recognition for her lead role as Erin Hardesty in the horror film The Texas Chainsaw Massacre (2003). Biel has since starred in such films as The Rules of Attraction (2002), Blade: Trinity (2004), Stealth (2005), The Illusionist (2006), I Now Pronounce You Chuck & Larry (2007), Valentine's Day (2010), The A-Team (2010), New Year's Eve (2011), Total Recall (2012), and Hitchcock (2012).

In 2017, Biel was the executive producer and star of the USA Network limited drama series The Sinner, for which she received nominations for a Golden Globe Award and a Primetime Emmy Award for Outstanding Lead Actress in a Limited Series or Movie.