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Tucci, an Italian American, was born in Peekskill, New York, the son of Joan (née Tropiano), a retired secretary and writer, and Stanley Tucci, Sr., a retired high school art teacher. His sister is actress Christine Tucci, and his cousin is the screenwriter Joseph Tropiano. He grew up in Katonah, New York and attended John Jay High School. Stan played primarily on the John Jay soccer team and enjoyed playing a couple of years on the John Jay baseball team, however, his main interest lied mainly in the school's drama club, where he and fellow actor and high school buddy, Campbell Scott, son of the late, great actor George C. Scott, gave well-received performances at many of John Jay's drama club productions.

Tucci made his Broadway debut in The Queen and the Rebels on 30 September 1982. His film debut was in Prizzi's Honor (1985). Tucci is known for his work in films such as Kiss of Death, Road to Perdition, and Big Night, and in the television series Murder One as the mysterious Richard Cross. Big Night (1996), which he co-wrote with his cousin Joseph Tropiano, starred in, and co-directed with Campbell Scott, premiered at the Sundance Film Festival. The film also featured his sister Christine and mother, who wrote a cookbook for the film. It won him and Tropiano the Independent Spirit Award for Best First Screenplay.

He has been nominated twice for Golden Globes, and won both times — for his title role in Winchell (1998), and for his supporting role as Adolph Eichmann in Conspiracy (2001), both for HBO films. He also received a Screen Actors Guild Award nomination for Winchell. He was nominated for Broadway's Tony Award as Best Actor in a Play for his role as Johnny in the 2002 revival of Terrence McNally's Frankie and Johnny in the Clair de Lune.

In July 2006, Tucci made an appearance on the USA Network TV series Monk, in a performance that earned him a 2007 Emmy for Outstanding Guest Actor - Comedy Series. Tucci's latest TV series, the medical drama 3 lbs., debuted on CBS in the 10:00 p.m. EDT time slot on November 14, 2006. It was cancelled on November 30, 2006 due to low ratings. Tucci is the voice of Cingular Wireless/AT&T. He can be heard as the voice over in their "Raising the Bar" campaign. In 2007, Tucci had a recurring role in medical drama ER. In 2009, Peter Jackson's adaptation of The Lovely Bones will hit the cinemas, in which Tucci portrays George Harvey, the murderer of the lead character (portrayed by Saoirse Ronan).

Tucci lives in Westchester County, New York with his wife, Kate, and their three children, twins Isabel and Nicolo, and Camilla. Tucci was co-owner of the Finch Tavern restaurant in Croton Falls, New York, which is now the Primavera Restaurant.