In an alternate universe, heroic analogues of Lex Luthor and the Joker (called the Jester) are attempting to steal a device from the headquarters of the Crime Syndicate. The pair trip an alarm but manage to secure the "Quantum Trigger". The Jester sacrifices himself to allow Luthor to escape and kills J'edd J'arkus and Angelique (alternate versions of Martian Manhunter and Hawkgirl respectively) with a radioactive bomb. Luthor is confronted by the remaining Syndicate members - Ultraman, Superwoman, Power Ring, Johnny Quick and Owlman, but escapes to the Earth of the heroic Justice League by activating a dimensional travel device.
Luthor locates a police station and is mistaken for the evil Luthor where he ends up strip-searched. The Justice League are summoned and Superman's x-ray vision confirms Luthor's reversed organs indicate that he is from a parallel Earth and that the evil Luthor is still incarcerated at Stryker's Island. The Justice League take the alternate Lex Luthor to their Watchtower, where the heroes learn of the Syndicate threat. As the Justice League debate the matter, Luthor hides the Quantum Trigger on the satellite. With the exception of Batman, who decides to stay as he believes the League is too busy with their own issues, the group - Superman, Wonder Woman, Green Lantern , Flash and Martian Manhunter - travel to Luthor's Earth.
Arriving at the Justice League's base on Luthor's parallel Earth, the heroes encounter and defeat two of the Syndicate and their allies (also evil versions of heroes belonging to the League). After another battle in the skies above the base, the heroes regroup at the Jester's headquarters and plan to attack Syndicate targets. After a successful series of raids and capturing Ultraman, the League confront United States President Slade Wilson. Wilson releases Ultraman and explains that acceding to the Syndicate's demands saves millions of lives. His daughter, Rose, however, regards him as a coward. J'onn inadvertently reads her mind and explains that as a military man her father actually holds life more dear than others. J'onn foils an assassination attempt on Rose and the pair fall in love.
Owlman constructs a weapon, the Quantum Eigenstate Device or Q.E.D., which the Syndicate intend to use as the equalizer to the threat of a nuclear reprisal. When pressed by Superwoman, Owlman admits the weapon can destroy entire worlds. Believing there are many parallel Earths, and that each one develops from the choices that each person makes, Owlman becomes obsessed with the idea that nothing he does can possibly matter, as there will always be parallel worlds where he explored another option. Owlman begins fervently seeking Earth-Prime, the very first Earth from which all other universes originated. Owlman intends to use the weapon to destroy Earth-Prime, causing a chain reaction that would erase the entire multiverse, as it is the only action that would not result in the creation of another universe. He then sends Superwoman with three of her lieutenants to the League's dimension, and on the Watchtower they battle Batman, Aquaman, Black Canary, Black Lightning, Firestorm and Red Tornado. Superwoman and one of her lieutenants escape with the Quantum Trigger, but are followed by Batman.
Batman tricks and defeats Superwoman, and summons the League. J'onn and Rose bond, and Rose decides to learn the location of the Syndicate base to allow the Justice League to confront them. The League arrive at the Crime Syndicate's moonbase with captive Superwoman, and eventually battle the Syndicate. Owlman fights off Batman and takes the QED bomb to Earth-Prime, the original universe that contained the first alternate Earth. Earth-Prime turns out to be uninhabited and lifeless, having suffered a cataclysm that caused it to leave solar orbit. Luthor speculates that a speedster might be able to vibrate and match the temporal vibration of the teleported QED device and open a portal. Batman dissuades the Flash from attempting this and Johnny Quick volunteers and opens a portal.
Batman pursues Owlman to Earth-Prime, and although Owlman's exoskeleton provides an edge he is ultimately tricked and teleported with the Q.E.D. device to another lifeless, uninhabited Earth covered in ice with an orbital ring that either never coalesced into the Earth's Moon, or the shattered remnants of a Moon that passed too close to the Earth's Roche limit in that universe. Owlman decides not to abort the bomb's countdown and states that "it doesn't matter". He is killed when the bomb detonates and destroys that alternate Earth but no others.
Batman returns to discover that the strain of acting as a vibratory conduit has aged Johnny Quick to near death. Before dying, Johnny correctly deduces that Batman had anticipated this and prevented the Flash from trying to save him. J'onn returns, accompanied by President Wilson and the U.S. Marines, and together they arrest Ultraman, Superwoman, and Power Ring.
President Wilson thanks the heroes, and although Rose asks J'onn to remain with her, the group return to their dimension. Wonder Woman retains the still-cloaked plane stolen from Owlman, and Batman and Superman discuss a membership drive, with the five heroes summoned previously greeting the League.