The trailer, which was carrying the monkeys to a laboratory in Florida, collided with a dump truck on a motorway in Pennsylvania.
No-one was hurt, but four monkeys managed to escape.
Three have since been found, but one monkey remained unaccounted for on Saturday morning.
The long-tailed macaques are in demand for coronavirus vaccine research.
Pennsylvania State Police Trooper Andrea Pelachick confirmed the hunt for the escaped monkeys in Montour County.
"The vehicle involved was following 100 monkeys, and when the crash occurred a handful of monkeys got out," she told NBC News.
Police later confirmed that one monkey remained on the loose, but urged people not to attempt to look for or capture the animal. The public have been asked to call 911 if they spot the animal.