Barry Bonds has hit his 756th home run, beating Hank Aaron's all-time record of 755 home runs in American Major League baseball, set in 1976.
The San Francisco Giants slugger broke the record in front of his home crowd, hitting the run off a pitch from Washington Nationals' Mike Bacsik.
His achievement has been overshadowed by claims he has used steroids.
He has never tested positive for drug use and denies knowingly taking performance-enhancing subtances.
Nevertheless, rumours have persisted since his home run tally took off after he turned 35, moving past the legendary Babe Ruth into second place in May last year with his 715th career home run.
"Thank you very much. I got to thank all of you, all the fans here in San Francisco. It's been fantastic," said Bonds, after he crossed home plate.
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"To my dad," he said, pointing to the sky. "Thank you for everything."
Hank Aaron was not present to see his record broken, but he sent a tape message of congratulation.
"It is a great accomplishment which required skill, longevity and determination," Aaron said.
The BBC's David Willis, in California, says that to fans of America's national sport no record means more than the home run record.
But away from the fireworks and celebration at the San Francisco Giants' stadium, a cloud hangs over his achievement, our correspondent says.
In September, 2003 US federal agents raided a nutritional supplements company in the suburbs of San Francisco.
The company, Balco, was found to be supplying performance enhancing drugs to top American athletes.
Among those arrested and subsequently jailed was Barry Bonds' trainer.
Bonds himself has always denied wrongdoing and has never failed a drugs test.
But, our correspondent says, even die-hard fans question whether his strength with the bat is for real.
No matter how reluctantly, many believe that in setting a new record in American sports Bonds has almost certainly cheated, he adds.
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