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3,000 years ago, warlord Yaotl enters a portal into a parallel universe and becomes immortal, but his four generals are turned to stone. The portal releases 13 immortal monsters that destroy his army and his enemies.
In the present, the Teenage Mutant Ninja Turtles have grown apart. After defeating the Shredder, Master Splinter has sent Leonardo to Central America for training. Donatello works as an IT specialist, Michelangelo works as a birthday party entertainer called ”Cowabunga Carl”, and Raphael works at night as the vigilante ”Nightwatcher”. April O’Neil operates a company that locates and acquires relics for collectors, assisted by her boyfriend, Casey Jones.
April tells Leonardo that the turtles have drifted apart. April returns to New York with a statue for wealthy tycoon Max Winters. Leo also returns, and April and Casey deliver the statue to Winters. Winters hires Karai and the Foot Clan to search the city for the 13 monsters before the portal opens again. Raphael encounters Casey, who reveals his knowledge of Raphael’s double identity and joins him in hunting criminals. Winters, who is actually Yaotl and is still immortal, reanimates his stone generals. Leo returns to the sewer, meeting Splinter. Splinter forbids the Turtles from fighting until they can act as a team again. While training, the Turtles encounter one of the 13 beasts battling the Foot Clan. The Turtles engage the beast, going against Leo and Splinter’s orders. When Raphael visits Casey, they encounter one of the monsters and witness its capture by the Foot and the Stone Generals, who spot them and Raphael is knocked unconscious. Casey takes Raphael back to the apartment while April calls the Turtles for help and reveals the identities of Yaotl and his generals. After being revived, Raphael suggests they pursue Yaotl, but Leo forbids him to go until Splinter gives out the order. Raphael investigates alone.
Leo, Donny and Mikey plan their next move, where Donny discovers the next portal will open over Winters’ tower. Splinter tells Leo that his team is incomplete, and that he knows what he must do. After 11 monsters have been captured, General Aguila questions Yaotl’s actions. The generals conspire to betray Yaotl, wanting to remain immortal. Raphael encounters one of the remaining monsters but drives it off. Leo fights Raph, but when he breaks Leo’s twin swords, Raphael is forced to leave. The generals seize Leo, intending to substitute him for the 13th missing beast, and Raphael decides to make amends by rescuing Leo. As the portal opens, Yaotl discovers his generals’ treachery, while Splinter and the Turtles, accompanied by Casey and April, fight their way through the Foot Clan cordon and breach the tower. Yaotl reveals the truth to the heroes: he wants to be free of his curse of immortality. The generals reveal that they wish to preserve their immortality, but also to use the portal to bring in more monsters to conquer the world.
Having refused to betray Yaotl in exchange for serving the Generals, April, Casey and the Foot Clan work together, searching for the final monster while the Turtles fight the generals. Splinter and Yaotl fight off numerous monsters emerging from the portal. April, Casey and Karai arrive at the tower with the last monster. The monster crashes into the Generals, dragging them into the portal before it closes. Karai warns them to enjoy their victory while it lasts, claiming they will soon contend with a familiar foe, which the Turtles suspect to be the Shredder. She and the rest of the Foot Clan depart. Yaotl, now mortal, honors the Turtles and Splinter, thanking them for fulfilling his wish before disappearing. Splinter places Yaotl’s helmet among his trophy collection, as well as Raphael’s ”Nightwatcher” helmet and Michelangelo’s turtle costume. Raphael says that the Turtles will always be brothers.