Plot

April O'Neil (Megan Fox) is a reporter for Channel 6 news in New York who has been researching a gang called the Foot Clan which has been terrorizing the city. She questions a dock worker about stolen chemicals, typically used in genetic research, that may be linked to the Foot Clan. Later that night, she returns and witnesses the Foot Clan unloading cargo. April tries to record footage using her phone, but a shadowy figure arrives and takes out the Foot Soldiers one by one. She tells her coworkers and her boss Bernadette Thompson, but no one believes her story. Meanwhile concerned by the deteriorating security situation in light of the Foot Clan attacks, the city of NY signs a lucrative deal with Eric Sacks of Sacks industries to provide security equipment to its crime fighting units. Eric Sacks reveals that many yrs ago his lab was attacked by the Foot Clan, which resulted in the death of many of his top research scientists (which included April's dad).

The surviving Foot Clan soldiers report the emergence of a vigilante to their leader, the Shredder, who instructs his second in commands to start terrorizing the city to make the vigilante(s) to reveal themselves.

The Foot Clan next attacks a subway station and hold hostages in order to lure the vigilante out. April rushes to the scene, hoping to encounter the vigilante or find evidence confirming his existence, but ends up getting held hostage by The Foot Clan. However, four figures arrive and defeat the Foot. April tracks the vigilantes to a rooftop and takes a picture of them, causing the vigilantes (humanoid mutant turtles Leonardo (Pete Ploszek), Michelangelo (Noel Fisher), Donatello (Jeremy Howard) and Raphael (Alan Ritchson)) to notice her and demand her to give them the camera. April then passes out at the sight of the turtles. When she comes to, she learns that the turtles deleted her photo and catches some of their names, finding them familiar. The turtles warn her not to divulge their existence before leaving, but she takes another photo of them.

Upon returning to their lair, the sewers, the turtles are caught by their master Splinter (Tony Shalhoub). Splinter scolds them for going outside to the city. He slaps them all across the face, then takes them to the "Hashi". April runs home and opens a box filled with documents, pictures, and July 1999 videos on "Project Renaissance" which involved her now-deceased father Dr. O'Neil. She notices that the turtles she cared for from her father's laboratory fifteen years earlier seem similar to the Ninja Turtles. Recalling the development of a mutagen by her father, she eventually realizes through research that the Ninja Turtles are the turtles from the laboratory.

Once again, she tries to convince Bernadette Thompson that the Turtles are real. Bernadette becomes so infuriated that she fires April. April tries telling her cameraman Vern Fenwick (Will Arnett) about them, but he does not believe her either. He does agree to take her to the home of her father's lab partner Eric Sacks (William Fichtner), a famous scientist and the CEO of Sacks Industries. April tells him about the Ninja Turtles, showing him the photo. Sacks explains Project Renaissance: he and her father were cultivating the mutagen for its healing properties that could repair organic material at a cellular level. Sacks theorizes that, when the laboratory was destroyed, the mutagen must have somehow caused the turtles to mutate into humanoids. Also, the mutants blood is now the last known source of mutagen as all of Eric's work was destroyed in the fire.

Meanwhile in the sewers, the Turtles are in the "Hashi" (which is an area located in the sewer where the turtles must stay in torturous positions until they reveal where they were) where the turtles remain silent about where they were that night. Splinter then eventually uses a 99-cheese pizza to force Michelangelo to reveal that April has spotted them, prompting Splinter to order them to find April and bring her to their lair (which is the sewer) as she is now in great danger from the Foot Clan since she made contact with the Turtles.

The Turtles find April and blindfold her, so she won't know their lair's location. They take her to Splinter, who explains how she saved their lives years before when she rescued them from the fire and released them into the sewers. As the turtles and Splinter grew more intelligent from the mutagen over the past 15 years, he started to teach the turtles how to defend themselves in the art of Ninjitsu from a martial arts book he found in the sewer. April admits that she has told her father's associate about them, unaware that Sacks is actually the adoptive son of the Foot Clan's leader called the Shredder (where Sacks created his armor). Splinter tells April about the real association between Eric & Shredder.

Meanwhile, Sacks relays the information to Shredder. Shredder and Sacks plan to spread a deadly virus throughout New York causing a quarantine in order to seize control by offering the mutagen as a cure. Shredder needs the Turtles to extract the mutagen from their blood. After the virus is spread all over New York, Sacks plans to sell the mutagen cure for a massive profit and making him even more rich. Sacks had the exact same plan 15 yrs ago, when April's dad found out about it & destroyed his work, getting killed in the process.

The Foot, accompanied by Shredder, find Splinter and the Turtles in the sewers and a battle ensues. They are overwhelmed and Shredder captures Leo, Donnie and Mikey and leaves Splinter severely injured. Thought dead, Raphael survived the wreckage of the lair. Splinter instructs Raphael and April to save the other three Turtles. April calls Vern to give them a ride to the laboratory where the other three Turtles are being held. When they arrive, April frees the Turtles, who join Raphael in fighting Shredder, but Shredder escapes. April, the Turtles, and Vern escape down a snowy mountain with The Foot and Karai in pursuit and manage to get away.

The Turtles plan to attack Shredder on the rooftop of Sacks' building before he is able to release the toxin, while April and Vern search for the mutagen and battle Sacks inside the building. He reveals to April that he killed her father for burning down the lab. As he closes in on April, Sacks is knocked out by Vern using a microscope. April finds the mutagen and heads onto the rooftop to give it to the Turtles. With April's help, the Turtles finally defeat Shredder who falls off the roof As he is immediately surrounded by the police, he touches the spilled mutagen and begins to grow larger.

That night, Vern attempts to impress April with a Ford Crown Victoria Police Interceptor. This unfortunately fails when the Turtles come in with a vehicle of their own and accidentally blow it up with an RPG. The turtles then offer April a ride home, but she kindly turns down the offer. The film ends with Mikey serenading April with "Happy Together" (a love song performed by 60's rock band The Turtles).