Mitt Romney was on the defensive after raising the discredited rumour that Barack Obama was not born in the United States and therefore ineligible to be president.
The Republican candidate jokingly declared "no one's ever asked to see my birth certificate" as he campaigned near his own Michigan birthplace.
Mr Romney later insisted the remark was just a joke and not meant to question Mr Obama's citizenship.
But the comment risked creating an unwanted distraction for Mr Romney in his last few days of campaigning before the Republican National Convention, which opens on Monday and concludes with his formal nomination later in the week.
The authenticity of Mr Obama's birth certificate has been questioned by some Republican critics who insist Mr Obama is not a "natural-born citizen" as required by the Constitution.
Mr Obama released a long-form version of his birth certificate last year as proof that he was born in Hawaii in 1961.
But polls show some Republicans remain unconvinced. A Pew Research Centre poll taken in April found 19% of Republicans, when asked where Mr Obama was born, said they were not sure, and 6% believe he was born in another country.
The flap over the joke came a day after Mr Romney caused another stir by declaring that big business was "doing fine" in the current struggling economy in part because companies get advantages from offshore tax havens.
Mr Romney made his birth certificate remark at a large outdoor rally in Michigan, where he grew up and where his father, George Romney, served as governor.
Mr Romney told supporters that he and his wife, Ann, had been born at nearby hospitals.
"No one's ever asked to see my birth certificate. They know that this is the place that we were born and raised," Mr Romney said.
The crowd of more than 7,000 responded with hearty laughter.