The gameplay is very similar to the arcade games of the Konami era, in which the Turtles are controlled by an 8-way joystick, and jump and attack buttons. A key addition to this game is Turtle Power. The meter fills up as the Turtles defeat enemies. Once it's full, the player can press the Turtle Power button to unleash a special attack that instantly defeats most enemies and does significant damage to boss enemies.
The starting three levels can be played in any order.
- Shredder's Hideout (Unlocked by completing the previous three levels)
- Boss 1: Bebop
- Boss 2: Rocksteady
- Boss 3: Chrome Dome
- Boss 4: Baxter Stockman
- Final Boss: Shredder
This game has a much greater variety of power-ups than the Konami era video games.
- Pizza: Replenishes about 50% of the health meter.
- Turtle Power Icon: Provides a small boost to the Turtle Power meter.
- Smoke Bomb: Can be thrown on the ground to temporarily stun all enemies.
- Shuriken: Gives the player 10 shuriken to throw at enemies.
- Shell Icon: Temporarily makes the Turtles spin in a small tornado, instantly defeating any enemy they touch, just like the bomb pizzas in Turtles in Time.
- Assist: Summons one of the Turtles' allies to attack all on-screen enemies.
- Leatherhead: Rolls across the floor to attack enemies.
- Metalhead: Shoots missiles at the enemies.
- Ice Cream Kitty: The Turtle throws her cooler to attack an enemy, then she changes direction. She leaves the screen after a short while.
- Donatello (Rob Paulsen)
- Turtle Power: Creates a ball of electricity in his hand, and another at the end of his bō, electrocuting enemies.
- Leonardo (Seth Green)
- Turtle Power: Spins and becomes a tornado, trapping enemies.
- Michelangelo (Greg Cipes)
- Turtle Power: Deflects raining pizzas with his nunchucks.
- Raphael (Sean Astin)
- Turtle Power: Repeatedly punches the ground while surrounded by a fiery aura.
- Baxter Stockman (Phil LaMarr)
- Bebop (J.B. Smoove)
- Chrome Dome (Nolan North)
- Kraang Subprime (Gilbert Gottfried)
- Krang (Pat Fraley)
- Rocksteady (Fred Tatasciore)
- Shredder (Kevin Michael Richardson)