The legendary outlaw, Ben Wade (Russell Crowe) and his gang have just robbed an armed stage coach carrying the Southern Pacific Railroads payroll. They kill everyone on-board except Pinkerton guard, Byron McElroy (Peter Fonda) whom Charlie Prince (Ben Foster), Wade's right hand man, shoots in the belly. A poor rancher Dan Evans (Christian Bale) and his two sons, William (Logan Lerman) and Mark (Benjamin Petry), find McElroy and take him to Bisbee, Arizona to find a doctor.
Ben Wade separates from his gang and goes to meet with lovely Emma Nelson (Vinessa Shaw) in a nearby saloon, but learning of his presence in town, the sheriff immediately surrounds the building. Dan distracts Ben Wade, who is captured without a shot being fired. Railroad representative Grayson Butterfield (Dallas Roberts) asks for paid volunteers to join McElroy's posse to take Wade to the train station in the town of Contention three days away. The train is due at 3:10, and they must put Wade on the train's prison car bound for Yuma. Once Wade arrives in Yuma, he will receive a quick trial in Federal Court and be hanged. Dan Evans, desperately needing money to save his farm, agrees to help transport Wade to Contention but he must be paid $200. The other paid volunteers who join the posse are Tucker (Kevin Durand) and Doc Potter (Alan Tudyk), the town's veterinarian. Unfortunately for the posse, Charlie Prince sees what has happened to Wade, and he goes after the rest of the gang to help liberate Wade.
Despite a deception to fool the gang, the odds move in their favour, and in the end it's Dan's desire to impress his sons that means getting him on the train becomes a personal battle of wills.
In a hotel in Contention City, where the remains of the posse hide out, Wade continues to bargain for his freedom. Despite very tempting offers, Evans ignores his pleas. Wade's gang rides into town, and, after locating their boss, recruit as many of the townspeople as possible with an offer of $200 for the successful killing of any member of Evans' posse. The sheriff and his men, previously offering help to Evans, decide to surrender. They are gunned down, and Butterfield, terrified, offers Evans the $200 to just walk away. Evans, realizing the parallels between his current situation and the one in which the government paid for his leg lost in the Civil War, says, "It's funny, if you think about it - which I have been lately - is...they weren't paying me to walk away. They were paying me so they could walk away." Butterfield promises to see Evans' son home, but Evans want Butterfield to promise more than that. Butterfield is convinced to pay off Evans' debts, make sure his water flows, and that he will give $1000 dollars to his wife on Butterfield's return. Butterfield agrees, and he and William hold up in a room down the hall.
When the time for action, Evans brings Wade outside and they run for the station. Wade's gang pursues them relentlessly, and they barely make it to the station in one piece. At the very moment that the protagonist Dan Evans is putting Ben Wade onto the prison train, Wade's gang shoots the rancher to death. Ben Wade takes his gun from his right-hand man Charlie Prince and, in an act of vengeance for the rancher who had earned his respect, murders every man of his gang. Ben Wade boards the prison train as a final act of symbolic tribute to the rancher who died getting him there, and whistles for his horse as the train drives into the distance.