The one-eyed bartender is clearly missing his right eye, but when he turns to his left and we see the right side of his face, his right eye is open. When he then he turns back towards the girls, his right eye is gone again.
During the attack in Hollywood, Lucy is able to burst Bratt's gum bubbles with her bare hands. However the Minions cannot burst the bubbles with battle-axes, power tools and pitchforks.
Bratt is right handed, and his keytar is also right-handed. Gru is left-handed, and when he steals the keytar at the end of the movie, it is now somehow also left-handed.
In an early scene Clive the Robot ejects a cassette tape and spins it and inserts it to play the other side, which would put the open tape end on the wrong side of the tape deck. It needs to be spun around as well to keep the open tape end on the correct side of the deck.
When the giant Bratt robot collapses, we can see the L of the Hollywood sign leaning against the O. As Gru climbs up the robot, the L is now suddenly upright.
At the beginning when Balthazar Bratt does his diamond heist, the AVL obtain incorrect coordinates of his location. His heist took place off the coast of Hollywood, however on an online map the coordinates show that he was in Tennessee.
During Gru and Dru's heist, Dru is climbing up the exterior walls of Bratt's lair with his special suit set on "Sticky" mode. He then has to switch to "Camouflage" when spotted by security systems. If he switched suit modes, then "Sticky" would have turned off, thus causing Dru to fall.
During the prison scene, Mel is doing tattoos and his mouth doesn't open when he exclaims about imprinting a tattoo on a Minion's butt.
The cop cars in the 80's TV show look too modern.
When Gru and Dru hook onto the side of the cliff, the hook doesn't hook at the very top. When they drove over the top and landed, that wouldn't have been possible as the hook didn't reach the top, and thus couldn't pull them all the way to the top.