However, after witnessing Eduardo's two-timing son, Antonio, sweep Margo off her feet, and invite everyone, including Margo, to his Cinco de Mayo party, Gru renews his focus on Eduardo.
The next day, the AVL arrest Floyd Eagle-san after an almost-empty jar of the PX-41 mutagen is found in his shop, and Ramsbottom declares the investigation closed.Lucy is reassigned to Australia, but before leaving she gives Gru her lipstick Taser as a reminder of her.Realizing that he genuinely loves Lucy, Gru is heartbroken, yet still cannot find the courage to ask her out on a date.Instead, he brings the girls to the Cinco de Mayo party and, whilst there, finds undeniable proof that Eduardo is indeed "El Macho".A mysterious vehicle using a huge magnet steals a highly potent mutagen known as PX-41 from a secret laboratory in the Arctic Circle.
The Anti-Villain League (AVL), headed by Silas Ramsbottom, tries to recruit former supervillain, Felonious Gru, to track down those responsible and recover the mutagen, but Gru refuses, claiming that he is now a legitimate businessman as well as the devoted father of Margo, Edith, and Agnes. Nefario, Gru's friend and assistant, tells Gru that he "misses being evil", and opts to leave him for new employment.
Gru reluctantly agrees to investigate the theft of the mutagen, and is partnered with undercover AVL agent Lucy Wilde.
The pair begin their search in the Paradise Shopping Mall, disguised as bakery employees.
Gru immediately comes to suspect Mexican restaurant owner, Eduardo Pérez, of being the infamous "El Macho", a legendary supervillain who supposedly died after skysurfing a shark into the center of an active volcano.
Gru and Lucy break into Eduardo's restaurant at night, but find nothing to prove that he is the culprit.
Meanwhile, Agnes confides her dream to one day have a mother, and believes that Gru will fall in love with Lucy, but, despite her excitement, Gru tells her that his relationship with Lucy is strictly professional.