Navratilova, dancing with partner Tony Dovolani, was booted from the popular television ballroom contest show after their jive dance on Monday received the lowest scores from the judges in the second week of competition.
Navratilova, 55, one of the greatest women's tennis players in history, was philosophical, saying "it was a great experience," although she said she wished she could have stayed longer.
"But I blew it yesterday, and it cost me," she admitted.
Two other bottom finishers based on the judges' ratings, Melissa Gilbert and Gladys Knight, were both declared safe from elimination early in the live episode. Viewer votes combined with judges' ratings decide which star is eliminated each week.
But one of the judges' top-rated competitors, Roshon Fegen, was among the dancers facing possible elimination when fans apparently failed to vote in line with the judges' raves about his quickstep on Monday night's performance episode.
The other bottom finisher with Fegen was musician Gavin DeGraw, who will live to dance another week.
The top scorers based on the judges' ratings after two weeks were classical singer Katherine Jenkins, Disney Channel star Fegen and telenovela actor William Levy.
Highlights of the Tuesday's elimination round included Matt Nathanson's performance of "Run" along with Sugarland, who also performed their hit "The Incredible Machine."
Audience members included Katie Couric, and last year's controversial contestant Chaz Bono, the transgender son of superstar Cher whose casting brought protests from some corners and an outpouring of support from others, including from his mother who attended several of his performances.
"Dancing With the Stars" annually is among the top-rated U.S. TV shows with a formula that pairs B-list celebrities, sports stars and singers with professional dancers performing sambas, waltzes and other dances across a ballroom floor.
(Editing by Chris Michaud)