When Woody, a pull-string cowboy doll, is stolen at a yard sale by a toy collector, his friends from Andy's room spring into action to rescue him. Woody discovers that he is a highly valuable collectible from a 1950s TV show called Woody's Roundup, and he meets the other prized toys from the show: Jessie the cowgirl, Bullseye the horse, and Stinky Pete the Prospector. While initially flattered by the attention, Woody is faced with a dilemma: he can choose a life of immortality as a museum piece or return to the loving, but finite, embrace of his owner, Andy.

The toy collector plans to sell Woody and the Roundup gang to a museum in Japan, making their separation from Andy and the rest of the toys seemingly inevitable. Meanwhile, back at Andy's, Buzz Lightyear leads a daring expedition to find Woody, recruiting friends like Mr. Potato Head, Slinky Dog, Rex, and Hamm for the mission. The group encounters numerous challenges and adventures along the way, including crossing treacherous city streets, navigating a toy store overrun by new Buzz Lightyear models, and facing an enemy from Buzz’s past. With time running out, Woody must decide where his loyalties truly lie, with the prospect of his own legacy and the impermanence of the world he's known looming over him.