Born to an Italian American father and a Polish American mother, Bello grew up in a working-class Roman Catholic family. She attended Villanova University, majoring in political science. She had every intention of becoming a lawyer, but took an acting class during her senior year, just for fun. She was soon cast in small off-Broadway plays, such as The Killer Inside Me, Small Town Gals With Big Problems and Urban Planning. She later guest-starred on episodes of The Commish (1991), Nowhere Man (1995), Misery Loves Company (1995) and Due South (1994).

Maria Bello as John McLoughlin's wife, Donna, in World Trade CenterBello got her breakthrough when producers Kenny Lenhart and John J. Sakmar cast her in the spy show Mr. & Mrs. Smith (1996) as Mrs. Smith; they remembered her from her performance in a failed pilot that was a remake of the classic TV series 77 Sunset Strip (1958/I). The show was cancelled after just eight weeks on the air. Then came a guest stint on ER as pediatrician Dr. Anna Del Amico, in which she guest-starred on the final three episodes of the third season. The show's producers were so impressed with her that they asked her back as a regular on the series. Bello also had a role in the movie Coyote Ugly. She has been twice nominated for a Golden Globe: In 2003, as Supporting Actress for The Cooler and in 2005, as Best Actress for A History of Violence.

Bello practices yoga and ballet. Bello and her TV executive boyfriend, Dan McDermott, welcomed their first child, Jackson Blue McDermott, on March 5, 2001. She is currently involved with actor/director Tony Goldwyn.