Haley, who had described Trump as unfit for office during her campaign, urged her supporters to vote for him over Democratic President Joe Biden "for the sake of our nation."
"You don't have to agree with Trump 100% of the time to vote for him," Haley, a former U.N. ambassador and South Carolina governor, said after taking the stage to a mixture of cheers and boos. "Take it from me."
DeSantis, the conservative Florida governor whose campaign sputtered early in the year, received a warm welcome from the crowd as he attacked Biden, 81, as too old for the job.
His right ear bandaged after Saturday's assassination attempt, Trump applauded from his box in the arena, where sat alongside running mate U.S. Senator J.D. Vance. Vance, himself a former fierce Trump critic who has become a staunch supporter, will headline the convention's third night on Wednesday.
The show of harmony was intended to contrast with Democrats, who have spent weeks mired in intraparty tensions over whether Biden should abandon his reelection bid after his halting June 27 debate performance against Trump, 78.
Many of the evening's speeches in Milwaukee - centered on the theme of law and order - were infused with Trump's anti-immigrant rhetoric, with speakers angrily denouncing Biden's southern border policies.
Kari Lake and Bernie Moreno, who are running in high-profile U.S. Senate races in Arizona and Ohio, respectively, and U.S. senators Ted Cruz of Texas and Tom Cotton of Arkansas all called the flow of migrants an "invasion."
While border crossings reached record highs during Biden's tenure, arrests dropped sharply in June after the president implemented a broad asylum ban.
Anne Fundner, a mother whose teenage son died from fentanyl poisoning, said she held Democrats responsible. The family of Rachel Morin, a Maryland woman who authorities say was raped and killed by a Salvadoran immigrant who had crossed the U.S.-Mexico border illegally several times, blamed Biden policies as well.
Trump has highlighted Morin's murder on the campaign trail, where he frequently demonizes migrants as violent criminals. Studies show immigrants do not commit crime at a higher rate than native-born Americans.
Trump has pledged to launch the largest deportation effort in U.S. history.
Some of the heated attacks contradicted the message of national unity Trump had promised to deliver after the attempt on his life at a Pennsylvania rally on Saturday.
But Lara Trump, his daughter-in-law and co-chair of the Republican National Committee, closed the night with a shift in tone, saying Americans should remember "there is more that unites us than divides us."
In the wake of the shooting, however, voter fears about the deeply polarized nation ahead of the Nov. 5 election seem only to have deepened.
Authorities have yet to identify a motive for the shooting. The 20-year-old gunman was killed at the scene by the U.S. Secret Service.
In his first campaign speech since the assassination attempt, Biden told Black voters in Las Vegas on Tuesday he was "all in" for reelection, again dismissing calls from some Democrats to step aside.
The four-day convention will culminate with Trump's prime-time address on Thursday, when he formally accepts the party's nomination to face Biden in a rematch of 2020.
