Winfield was born in Akron, Ohio. He attended Garfield High School in his hometown of Akron.
Winfield received an athletic scholarship to attend Ohio State University, where he played for the Ohio State Buckeyes football team from 1995 to 1998. As a senior in 1998, he was a consensus first-team All-American, and won the Jim Thorpe Award as the nation's top defensive back.
The Buffalo Bills selected Winfield in the first round (23rd overall pick) of the 1999 NFL Draft, and he played for the Bills from 1999 to 2003.
As a rookie, Winfield played in all 16 games. In the season opener, a 31-14 loss to the Indianapolis Colts, Winfield intercepted a pass from Peyton Manning and returned the interception for 3 yards. In Week 4 (October 4), Winfield recorded five special teams tackles in a 23-18 victory. Winfield got his second interception of the season in the Week 10 (November 14) 23-3 win over Miami with a 10-yard return. After Ken Irvin suffered an injury during the Week 15 (December 19) game against the Arizona Cardinals, Winfield took over and recorded three tackles and three hurries. For the final two regular season games of 1999, Winfield started for Irvin and perform 13 solo tackles in the Week 17 game against Indianapolis, the last game of the regular season. As a starter in the 2000 AFC Wild Card game, a 22-16 loss to eventual Super Bowl XXXIV runners-up Tennessee Titans and the Bills' last playoff appearance as of the 2013 season, Winfield recorded 6 tackles, intercepted one pass from Tennessee Titans quarterback Steve McNair, and returned the interception for 8 yards.
Winfield started all 16 games for the first time in 2001. In 2003, Winfield had 107 tackles, the highest single-season total of his career.
As a free agent, Winfield signed a six-year deal with the Minnesota Vikings on March 5, 2004.
In the Vikings' 30-27 win over the New Orleans Saints on Week 5 (October 6) of 2008, a Monday Night Football game, Winfield returned a blocked field goal attempt by Martin Gramatica 59 yards for a touchdown. Additionally, Winfield had 8 tackles and one deflected pass that game. Winfield finished 2008 with 95 total tackles, 2 sacks, 5 passes defended, and one interception.
On July 23, 2009 the Vikings signed Winfield to a new five-year, $36 million contract. The deal includes $16.1 million in guarantees. He made his second straight Pro Bowl in 2010 after replacing injured Dominique Rodgers-Cromartie. After making the 2010 Pro Bowl squad, he made an interception that led to a touchdown in an NFC 55-41 victory. He is regarded as one of the most physical corners in the league, known for his bone-jarring hits, and was ranked by Peter King of Sports Illustrated as the 31st best player in the NFL. He made his first Pro Bowl in 2008.
Winfield was the only player on the Vikings' active 2012 roster who played during the 1990s.
Winfield was released by the Minnesota Vikings on March 12, 2013 in a move to clear $7.25 million in salary cap space.
Winfield signed with the Seattle Seahawks on April 17, 2013 to a one-year, $3 million contract. He was assigned number 21 due to starting fullback Michael Robinson having already been assigned 26 in 2010. He was released from the team during final cuts on August 31, 2013.
Seattle Seahawks coach Pete Carroll announced on Twitter on August 31, 2013 Winfield was retiring.
His son, Antoine Winfield Jr., is a defensive back for the University of Minnesota. He was named to the ESPN true freshman All-American team in 2016.