The 39-year-old American, who made a comeback to the sport two years ago, said he was quitting for good to spend more time with his family and his charities.
Armstrong initially retired from cycling in 2005 but returned in 2009, finishing third in the Tour in his first year back. His final race was at the relatively low-key Tour Down Under in Australia last month.
"He's leaving a real legacy and to come back and get third in the Tour at the age he was and after what he'd already accomplished is pretty impressive," Armstrong's former team mate Christian Vande Velde told Reuters from the Tour of Oman.
Belgian cyclist Eddy Merckx, a five time winner of the Tour de France, said Armstrong was right to call it a day.
"It was time for him to stop," Merckx told Reuters from Oman. "He's won everything, had nothing left to prove to anybody."
Armstrong was diagnosed with testicular cancer in 1996 but survived and returned to the bike and went on to become one of the most successful and controversial cyclists of all time.
He won the Tour de France for an unprecedented seven consecutive years, from 1999 to 2005, before quitting the sport at the top.
He made a comeback in 2009 at age 37, saying he partly missed the thrill of competition but was driven by a greater cause, to help promote cancer awareness through his charity Livestrong.
Armstrong enjoyed mixed success but did not add to his record number of Tour de France wins. He finished a creditable third behind his team mate Alberto Contador in 2009 then 23rd last year, after moving to the RadioShack team but suffering a series of crashes that ruined his chances.
Armstrong announced late last year that January's Tour Down Under would be his last international race. He planned to compete in a handful of events in the U.S. this year, including the Tour of California, before revealing on Wednesday that he was quitting for good.
"My focus now is raising my five children, promoting the mission of Livestrong, and growing entrepreneurial ventures with our great corporate partners in the fight against cancer," he said.
Although he never failed a dope test and always denied doping, Armstrong was dogged by accusations of wrongdoing during his career.
Last year, former team mate Floyd Landis, while confessing to cheating himself, accused Armstrong of using performance enhancing drugs.
U.S. federal authorities are investigating Landis's claims, but have not laid any charges and Armstrong said he had nothing to fear because he had done nothing wrong.
"They can keep looking," he told reporters in Australia last month. "If you're trying to hide something, you wouldn't keep getting away with it for 10 years. Nobody is that clever."
(Additional reporting by Alasdair Fotheringham in Oman; Editing by Ken Ferris/Kevin Fylan)