Before the "royal huddle" decides to send Flick away, if you look closely at Princess Atta's mouth when she says "He can't mess anything up" you'll notice that the word "screw" is what was animated as opposed to the word "mess".
When Flick is leaving Ant Island, the two boys who follow him are making fun of him. One boy says, "My dad says he is going to die." When Flick left the Island, the same boy says, "Your dad is right. He's gonna die."
Flik couldn't reach the back of his invention when he tried to get a seed for Dot. Later, he is caught by a blade of grass and is flung backwards, and all the seeds fall out of his invention. Then we see him running toward the offering stone with his invention full of seeds. He couldn't have put them back in, because he can't reach his invention when it is on. How did the seeds get in there?
As Hopper is flying away carrying Flick, Tuck & Roll pull off his right antenna. Just as Hopper ditches Tuck & Roll, you see him flying away with both antennae still intact. Then scene flashes to Tuck & Roll holding removed antenna.
About 20 minutes into the movie, in Rosie and Dim's "lion tamer" scene, we clearly see that Rosie accidentally whips Dim's left foot. A few minutes later, as she's getting a bandage for him, he's whimpering and holding up his right foot for her.
At the very beginning, the sun is glinting off the water to the left of center, but when the view pulls back to show the island the shadows are all falling to the left, as if the sun is to the right of center.
When Dot flies after the circus caravan, and gets Flik to return, Flik is sitting off the back of the cart, so he can catch Dot. As soon as Flik turns down Dot's request for help, he's sitting in the front of the wagon.
Many of the characters have fewer legs than their real counterparts. Francis, Gypsy Moth, Manny, PT Flea, the mosquitoes, fireflies, and all the ants have four legs instead of six. Tuck and Roll have eight legs instead of fourteen. Heimlich has four thoracic legs instead of six. (The number of caterpillar prolegs can vary by species.)