Dos Santos tipped the scale at 240 pounds. Shortly thereafter, Velasquez weighed in at 241.
The Saturday night showdown will be the third between Velasquez and dos Santos. Dos Santos lifted the title from Velasquez in November 2011 with a first-round knockout. Velasquez regained the belt during a December 2012 rematch by dominating dos Santos en route to a lopsided unanimous decision.
"He's a tough dude," Velasquez said. "I'm ready for a tough fight. Whether it ends early or goes five hard rounds, I will be ready for whatever."
The usually smiling dos Santos had a stern look on his face from the moment he walked onto the stage to weigh in. He and Velasquez looked directly into each others' eyes during the traditional stare-down.
"It's going to be a great fight for the fans," dos Santos said. "I'm going to give them another knockout."
Daniel Cormier, who is ranked third among heavyweights by ESPN.com, weighed 224 pounds -- the lightest of his pro career. This fight is likely Cormier's last at heavyweight as he intends to compete at light heavyweight in his next cage appearance.
His opponent, Roy Nelson, came in at 249, making the co-main event official.