Amy Schumer’s comedy may not be as well known in Tampa Bay, Florida, as it is in New York.
When Ms Schumer called Donald Trump an “orange, sexual-assaulting, fake-college-starting monster”, several dozen people reportedly booed and walked out the venue.
The comedian introduced politics about half-way through the show, according to the Tampa Bay Times.
The audience members at the venue, which was used for the 2012 Republican National Convention, were asked to join her on stage if they supported the party nominee.
One man joined her but said his support was less about Mr Trump and more about his distrust of Hillary Clinton.
When some people booed, Ms Schumer asked them to leave and for security staff to remove anyone booing.
She told the audience how she had recently revealed that she was a victim of sexual assault.
One audience member, Marshal Knight, wrote on Instagram that Ms Schumer asked all sexual assault survivors in the audience to stand up.
“Powerful moment and DISGUSTED at the walkouts,” he wrote.
The comedian has been openly political in the spotlight, campaigning for stricter gun laws alongside her cousin, senator Chuck Schumer, since last summer when two women were killed in Louisiana during a screening of her film Trainwreck after a man opened fire.
She announced her support for Hillary Clinton in March and has attended campaign events in Florida to ensure people register to vote.
