Mitts, a member of the 2004 Olympic gold-medal team and one of the most recognizable players on the women's team, will be sidelined 6-8 months.
She had an MRI on Monday, and it confirmed the ACL tear. She will have surgery within the next 10 days.
"It is obviously very sad," said Mitts. "I was definitely feeling good about my game and how much I've progressed as a player over the past few years and ready to make a contribution to the team in the Women's World Cup. To have a setback like this is difficult to handle, but with the support of my family, my teammates, our medical staff and the coaches, I know I can come back strong. This team has become very special to me. I'm confident my teammates will bring home the World Cup, I just hope to be with them to bring home the gold in Beijing in 2008."
Mitts has played 71 times for the USA and worked her way in the starting lineup as an outside back. In the 6-2 win over Canada, she assisted on the second goal, the first of her international career, before going down with the injury.
"Heather has played great for us in 2007," said U.S. head coach Greg Ryan. "She put in one of the best crosses of the year on our second goal against Canada last Saturday night and we will definitely miss her at the outside back as we move forward in preparing for the Women's World Cup."
Mitts has done TV work as a commentator for ESPN on college football and Major League Soccer. She was voted espn.com's Hottest Female Athlete of 2004.