A gang of Jokerz launch a raid on Gotham Shipping to steal a large piece of electronic technology with three forklifts. The robbery is interrupted by the arrival of Batman. A fight ensues, which turns into a pursuit in the skies of Gotham City. One of the forklifts the Jokerz are using to carry away the equipment, piloted by Bonk, takes off to fight Batman, leaving the equipment and the other craft to crash. Batman overcomes Bonk, and his craft crashes. The equipment crashes to the ground, but Chucko manages to retrieve its memory chip and escape with the others before the equipment explodes.
Terry is puzzled as to why the Jokerz would be stealing advanced technology. Bruce Wayne theorizes that they're engaged in industrial espionage. Just then, Terry sees a news report that Wayne has announced his return to active leadership of Wayne Enterprises, to the disappointment of acting Operations Manager and rival-for-corporate-leadership Jordan Pryce. Terry resolves to wait for the Jokerz to make their next move. However, he does end up feeling the after-effects of a bite given to him by the spliced Jokerz member Woof in the earlier fight. However, he decides to go on his date with Dana Tan that evening, despite both the fact that such an injury would be difficult to explain to Dana, and Bruce's advice that he get a good night's rest instead. Bruce's advice ends up revealing itself to be quite valid, as Terry ended up so tired from the fight and injury that he not only fell asleep at the dance club, but seemed to be dozing through the dances and even talked to the wrong girl by mistake. Dana was initially annoyed, but soon pulled Terry off the dance floor to sit down and talk over coffee instead.
The Jokerz return to their lair, an abandoned candy factory. Chucko offers the memory chip, explaining that it was all he could get away with when Batman interfered. Their leader, hidden in the shadows, angrily smashes the chip with his fist and proceeds to berate them for their incompetence. Bonk is angered by this, and tells his fellow gang members that not only is their new leader is having them steal a lot of high tech equipment instead of money, but also that he never even explained to them what he planned to do with the components that they stole for them if he even had one. This fact made the Jokerz think it over and then glare angrily at their leader. When Bonk shouts that he wants out, the leader responds by pulling a gun on Bonk. Terrified, Bonk takes back his statements, but the leader pulls the trigger ― and a comical "BANG!" flag pops out. Bonk breathes a sigh of relief ― and the leader pulls the trigger again, shooting the flag into Bonk's chest like a dart, whereupon his bloodstream is quickly flooded with Joker venom. The leader steps out of the shadows, revealing himself to be the original Batman's old archenemy, the Joker. He now sports a skin tight purple outfit, short slicked-back hair, and is oddly lacking in eyebrows. The terrified remaining Jokerz swear their allegiance to him, then Ghoul begins searching the net for places to acquire a new piece of equipment while Dee Dee takes out the trash (by disposing of Bonk's eerily smiling corpse).
Bruce and Terry arrive at an evening gala at which Bruce plans to inaugurate his return. Said speech is immediately interrupted by familiar hysterical laughter. The spliced half-hyena Joker Woof then jumps into the crowd of spectators. During the distraction, Chucko and Ghoul break into the Wayne Enterprises lab downstairs and steal the desired technology, as well as gassing security. Up above, Bruce easily overcomes Woof with his cane. After checking on him, Terry dashes off to suit up. However, Wayne is then attacked by the newly arrived Dee Dee twins and is down on the ground when the Joker makes a dramatic emergence onto the stage, much to Bruce's shock and horror.
The now-costumed Terry returns, but is occupied with Woof as Joker and Dee Dee board a hovercraft. When Terry tries to pursue, Joker detonates a series of explosives, causing further chaos. Batman is forced to save civilians rather than pursue.
On the way back to Wayne Manor, Terry muses aloud that the Joker seems pretty vigorous for a man who should be in his mid-80's. A grim-faced Bruce simply orders him to carry on driving.
In another part of Gotham, the now middle-aged Tim Drake sees a news report on the Joker's re-appearance and is horrified.
Undaunted by Bruce's aloofness, Terry tries Commissioner Barbara Gordon for information, and receives much the same reply from her: That it isn't his business and that he shouldn't concern himself with it. As Terry leaves, he passes Drake, who has his own appointment with Barbara.
In the Batcave, Bruce runs a voice comparison between the Joker at the ceremony and his old recordings: They are identical. Terry returns, and realizes that the Joker, whom Bruce never mentioned once, must have been the greatest ever of Bruce's old foes. He asks how it's possible that Joker could still be alive, and Bruce says it's impossible: The Joker died years ago, and Bruce watched it happen.
Terry concludes that Joker was planning a crime so heinous that Bruce had no choice but to kill him. Bruce does not respond, but tells Terry that he wants him to hand in the Batsuit, reasoning that since Terry avenged his father's death, he has no need to continue wearing the cowl. Bruce thanks him for upholding the reputation of Batman, but declares that he was wrong to "force this life" upon Terry. Terry refuses to give up the suit, saying that fighting crime as Batman is what makes him a worthwhile person, after so many years as an irresponsible, lawless teenager. Bruce listens quietly, but turns away, calling Terry a "stupid kid" and claiming that he doesn't know what he wants, much like his previous sidekicks. A stunned and angered Terry retorts by throwing the Batsuit at Bruce's feet and leaving.
The next morning, Terry is having breakfast and reveals to his mother and Matt that he is no longer working for Mr. Wayne. Later on that night, Dana, Chelsea and Blade attend a club to have a girls night out, where Chelsea tries to persuade Dana to leave Terry and find someone new. However, to their surprise, Terry appears to them and leads Dana to dance, where he informs her that he no longer has his job, but tells her that they will finally get to spend more time together, much to her joy. However, as Terry and Dana are dancing, they are approached by two girls who drag Terry away from Dana and while they distract him, Ghoul suddenly appears and kidnaps Dana as Terry breaks away from the girls and begins to search for Dana, unknown to Terry the two girls are actually Dee Dee in disguise and begin to follow him.
At the same time, Bruce is in the Batcave where he is creating the antidote to Joker venom, suddenly Ace begins to bark which makes Bruce realize that someone has entered the Manor. Ace rushes out of the cave to confront the intruder, but becomes hurt. Bruce becomes concerned for Ace and goes to check on him but a gas bomb appears, as Bruce is hiding the antidote, the Joker appears before him and begins to attack Bruce. Back at the nightclub, Terry is still searching for Dana, but is soon attacked by the Jokerz. Chucko doesn't know why they've been sent to attack an apparent nobody, but as long as they have fun with it, they don't mind. As Terry fights them off, Dana breaks free from Ghoul and calls out to Terry for help as Ghoul recaptures her. Terry fights his way to Dana, but Ghoul throws her aside and as Terry rescues her she falls unconscious. Terry manages to send the Jokerz running, but Dana is sent off the hospital as she became injured during the fight.
Realizing that the Jokerz' boss must know his secret identity, calls Bruce in an attempt to get answers. However, when he gets Bruce's answering machine, Terry rushes to Wayne Manor, realizing that Bruce not picking up at night meant something bad must have happened.
Terry arrives at the Manor and sees that it has been badly vandalised. He finds a hurt Ace near the entrance to the Batave. Terry enters the Batcave to find the words "Ha! Ha!! Ha!" scribbled across the floor of the Batcave in red, mirroring how his father's place looked when he was seemingly killed by Jokerz. Horrified, Terry immediately runs down the stairs to find Bruce slumped against the computer, slowly laughing himself to death from Joker venom. Bruce manages to direct Terry to an overlooked vial of anti-venom lying under the chem table. Terry grabs it and and injects Bruce, whose rictus grin relaxes as the exhausted man falls unconscious.
Unsure of who else to turn to, Terry calls Barbara in for help. As she ministers to a shallowly breathing Bruce, Terry demands answers as he is a part of this conflict and therefore deserves them, Barbara reluctantly gives in and tells Terry the true story:
Some forty years earlier, after Bruce's old friend and partner, Dick Grayson, Nightwing had left to establish himself in another city, the second Robin, Tim Drake, is kidnapped by Harley Quinn and the Joker while he was out patrolling alone. After three weeks, Joker sends Batman and Batgirl a clue that leads them to the place where Tim is being held: the original Arkham Asylum (the old building had been partially demolished and the asylum had been moved to a newer high security building).
Upon their arrival, the Joker explains that he and Harley were "getting on" in years, and it is time to think about a family and they can't go through the joy of childbirth or do the adoption. And since Batman "always had a few spare kids" tagging along, the Joker "borrowed" one of them. He then reveals Tim, who has been brainwashed and disfigured into a miniature version of himself to make him their own son, much to Batman and Batgirl's horror. Joker takes off with Batman in pursuit, while Barbara fights Harley through the ruins of Arkham. In the fight, Harley falls off a cliff into a pit and vanishes.
Arriving in the asylum's old operating theater, which Joker has converted into a cinema, Batman sees "home movies" of how Joker tortured and brainwashed Tim. From his hiding place, Joker narrates the story, saying that Tim held out for a long time but finally broke down and revealed all of Batman's secrets, and delivers the final bombshell by calling Batman "Bruce".
Finding his hiding place, Batman engages Joker in a brutal fight and is ready to kill him but Joker manages to get the upper hand, disabling Bruce with a knife in his leg. As Bruce lies helpless, Joker tosses a gun to Tim, telling him to "deliver the punchline" and finish Batman. Batman softly calls Tim's name, and his young partner hesitates. After a moment of warring with himself, Tim shoots Joker fatally. As Barbara runs in, Tim suffers a complete nervous breakdown, drops to his knees and begins to cry. Batman limps over as Barbara gathers Tim into her arms to comfort him.
Bruce and Barbara bury the Joker deep under Arkham and take Tim to Leslie Thompkins, who helps him back to sanity over the course of the next year. The only other people who know what happened are James Gordon, Barbara's father who helped cover the incident up for the sake of his friendship with Batman, (and apparently Bruce's butler Alfred Pennyworth and Dick Grayson.)
Barbara sadly notes that with his final act of cruelty, Joker had in many ways gotten the last laugh after all -- and the physical and mental wounds from that time have never completely healed. Terry asks what happened to Harley after her fall. Barbara tells him that her body was never found, but even if she did survive, it's doubtful she'd be stirring up trouble after all these years. She also reveals to Terry that after Tim recovered, Bruce forbade him from ever being Robin again, blaming himself for what happened and swore that he would never endanger another young partner. Tim left and eventually married, fathered two children, and re-started his career as a communications engineer. Terry feels sorry for Tim after hearing the torture that he went through, but says Tim is still the most likely connection to the now-returned Joker. As Batman, he questions Drake, who denies any involvement and expresses resentment about his past life as Robin: all of them ― Barbara, Dick, and himself ― gave it everything they had, but it was never enough for Bruce's impossible standards. Tim even had a vain hope that one day he would succeed Bruce as Batman, but eventually grew sick of his life to the point that he detests even looking at his old Robin suit.
Finding Drake a dead-end, Terry then suspects Jordan Pryce. However, Terry finds both Pryce and the Jokerz on the Wayne Enterprises yacht and proceeds to eavesdrop. After the Jokerz try to kill Pryce, he rescues the man before a giant laser beam from a satellite destroys the ship. Although Pryce is not the Joker, Batman plays a recording of the conversation between Pryce and the Jokerz for two police officers nearby, revealing that Pryce has been helping the Jokerz steal the technology in exchange for them trying to kill Wayne. Pryce is arrested.
Terry returns to the Batcave, frustrated with his inability to solve the case. As he muses aloud to Ace, Bruce appears in his bathrobe, having partially recovered from the toxin. He confesses that this is why he didn't want Terry going up against the Joker, imposter or not, fearful that something similar would happen to him as it did with Tim all those years ago.
Looking around the Batcave, Terry notices something odd: even though Joker smashed the costume displays, only the Robin costume was deliberately damaged during his attack. Remembering Tim's resentment, Terry deduces that he may be behind this new Joker. Bruce is hesitant to consider Tim a suspect again, but Terry shows him on the Batcomputer how the stolen technology components, when utilized by a communications expert like Tim, could form a transmitter allowing him to hijack a military defense satellite — the same kind that destroyed the yacht. Somewhat chipper at the promise of a new lead, Bruce orders Terry to go after Tim and begins typing away at the computer. Terry remarks that he would also like to bring Ace along this time.
Terry tries to confront Tim at his job site, but discovers that he's been tricked by a hologram. Laser traps seal him in and motion-detecting blasters are mounted on the walls. Joker appears on a video screen, revealing that Tim has indeed joined forces with him. With his help, the Joker has taken control of an Hyperion class defense satellite, which he intends to use to bring Gotham under his control.
Terry manages to escape the booby traps set at Tim's lab and takes off in the Batmobile, being chased through Gotham by another giant laser beam. He tracks the Joker to the abandoned Jolly Jack Candy Factory. He is attacked by the Jokerz, but fights them off easily. One exception is Woof, but this time, Terry has brought along an equalizer, and Woof discovers that an encounter with Ace the Batdog is a rather painful experience.
Inside the factory, Terry finds an unconscious Drake. Awakening, Drake appears to be re-living the trauma of his kidnapping and how he killed Joker. Bruce, able to hear and see what's going on via the suit's visor and mic, is visibly saddened by Tim's words. Terry is also concerned, but then Drake turns on him, disabling him with an electrical device. To his and Bruce's horror, Drake then bursts into hysterical laughter, and transforms into the Joker.
The Joker reveals the truth behind his resurrection: using genetics technology that he stole years ago, he copied his consciousness and DNA onto a microchip and implanted it into Tim Drake's brain while Tim was his prisoner. He had to remain dormant inside Tim's brain for most of the past decades, but has been gradually getting stronger. Whenever the chip takes control of Tim, he transforms into the Joker. He then demonstrates that he can use Tim's voice to lie to his family should they grow worried. Soon, the Joker intends to take complete control of Tim's body.
The Joker prepares a satellite strike on Gotham, explaining that he plans to begin with either Wayne Manor, Dana's hospital, or Terry's house (by way of carving a smiley face on the city with the satellite's cannon), in order to crush Terry's spirit. But before he can fire, Terry sics Ace on him and uses the distraction to destroy the beam's guidance system. The beam then heads toward the factory.
The Joker declares the game over for the time being and attempts to escape, but Terry seals the factory. They face off in a final confrontation. This incarnation of the Joker is a much tougher physical combatant than his old self and soon shows himself to be more than a match for the young Batman, as having access to Tim's memories gives him knowledge of Batman and Robin's skills. As Joker gloats, Terry retorts that Joker doesn't know a thing about him, and proceeds to prove it by fighting dirty and catching the Joker off guard, landing a few good hits on him.
Terry soon changes his battle plan after Bruce informs him that Joker likes to talk, thinking that he does as well: hiding in the rafters, he plays mind games with the Joker, taunting him about his obsession with Batman and his sloppiness as a comedian, insisting, like the Joker told Bruce decades ago, that he's "pathetic," imitating the Joker's laugh at the ridiculousness of the "Clown Prince of Crime", saying that isn't funny of frightening, the psychological attacks which Joker has never suffered from the previously silent Batman sending his opponent into a crazed rage. An angry Joker throws a handful of grenades at his opponent, sending Terry crashing to the floor. The Joker pins him to the ground, pulls off his mask and begins choking him. Terry capitalizes on the Joker's distraction and destroys the microchip with Joker's own electrical joy buzzer, thus destroying the Joker once and for all and restoring Tim. Terry escapes with Tim and Ace before the satellite destroys the factory, taking the satellite jammer with it and killing the beam. The satellite then drifts off into space.
In the city jail, the Dee Dee twins are revealed to be the twin granddaughters of the aging Harley Quinn, who is seen bailing them out while lamenting what disappointments they are.
Barbara visits Tim in the hospital, assuring him that she's covering up for the Joker's crimes, of which he is completely innocent. Terry also comes to visit, and they are all surprised to see Bruce there. The Dark Knight explains that it's where he should be. The three old heroes mend their fences. Terry leaves, but not before both Tim and Bruce stop to tell him that he is truly worthy of the Mantle Of The Bat, and that Terry was wrong: it wasn't Batman that made Terry worthwhile. It was the other way around. Though neither expresses it overtly, both are respectful and moved.
The next night, Terry stands at the top of a high building, looking out over the city he protects. Like Bruce before him, this is the life he has chosen. He pulls on his mask, spreads his wings, and flies off into the heart of Gotham City.
- Ash Ketchum, Pikachu, Misty, Togepi, Brock, Molly Hale, Littlefoot, Cera, Ducky, Petrie, Spike, Scooby-Doo, Shaggy Rogers, Fred Jones, Daphne Blake, Velma Dinkley, The Sailor Scouts, Masters of the 100 Acre Team, The Human Mane 5, Lincoln and his 10 sisters, D-Team, Alpha Gang and their Dinosaurs, The Tenkai Knights (Guren, Ceylan, Chooki, Toxsa, Gen and Beni), The Bakugan Battle Brawlers and their Bakugan, Jack Skellington, Oogie Boogie, The Grand Duke of Owls, Kurumi Tokisaki, The Dazzlings, Bowser, Mistress Nine, Bowser Jr., Granox, Slyger, Attila and Hun, Oakley and Annie, Cassidy, Butch, Dr. Facilier, Grizzle, Spectral Space Pirates, Darla Dimple, Broccoli Alien Overlord, and Team Rocket guest stars in this film.
- Oogie Boogie, The Grand Duke of Owls, Kurumi Tokisaki, The Dazzlings, Bowser, Mistress Nine, Bowser Jr., Granox, Slyger, The Fearsome Five, Attila and Hun, Oakley and Annie, Cassidy, Butch, Dr. Facilier, Grizzle, Spectral Space Pirates, Darla Dimple, Broccoli Alien Overlord, and Team Rocket will work for the Joker in this film.
- Jason Marsden as Tino Tonitini
- Grey DeLisle as Lor McQuarrie and Daphne Blake
- Phil LaMarr as Carver Descartes
- Kath Soucie as Tish Katsufrakis
- Rebecca Shoichet as Sunset Shimmer
- Carly McKillip as Sakura Kinomoto
- Josh Grelle as Shido
- Michelle Rojas as Tohka
- Bryn Apprill as Kotori
- Tia Ballard as Yoshino and Yoshinon
- Michelle Lee as Origami
- Felecia Angelle as Mana
- Veronica Taylor as Ash Ketchum and Hun
- Ikue Ōtani as Pikachu
- Rachael Lillis as Misty and Jessie
- Satomi Kōrogi as Togepi
- Eric Stuart as Brock, Butch, and James
- Megan Hollingshead as Cassidy and Annie
- Marc Thompson as Attila
- Scott McAfee as Littlefoot
- Candace Hutson as Cera
- Heather Hogan as Ducky
- Jeff Bennett as Petrie
- Rob Paulsen as Spike
- Ashleigh Ball as Rainbow Dash and Applejack
- Andrea Libman as Pinkie Pie and Fluttershy
- Tabitha St. Germain as Rarity
- Cathy Weseluck as Spike the Dog
- Frank Welker as Scooby-Doo, Fred Jones, Woof the Hyena-Man and Ace the Bat-Hound
- Matthew Lillard as Shaggy Rogers
- Mindy Cohn as Velma Dinkley
- Terri Hawkes as Serena/Sailor Moon
- Karen Bernstein as Amy/Sailor Mercury
- Katie Griffin as Raye/Sailor Mars
- Susan Roman as Lita/Sailor Jupiter
- Stephanie Morganstern as Mina/Sailor Venus
- Lisa Ortiz as Oakley
- Kazumi Evans/Wubcake as Adagio Dazzle
- Maryke Hendrikse/Wubcake as Sonata Dusk
- Diana Kaarina/Wubcake as Aria Blaze
- Will Friedle as Terry McGinnis/Batman II
- Kevin Conroy as Bruce Wayne/Batman
- Mark Hamill as The Joker and Jordan Price
- Angie Harmon as Police Commissioner Barbara Gordon
- Tara Strong as Sci-Twi, Barbara Gordon/Batgirl (young)
- Dean Stockwell as Tim Drake
- Mathew Valencia as Tim Drake/Robin (young)
- Teri Garr as Mary McGinnis
- Arleen Sorkin as Harley Quinn
- Melissa Joan Hart as Delia & Deidre Dennis/Dee-Dee
- Don Harvey as Charles Buntz/Chucko
- Michael Rosenbaum as Stewart Carter Winthrop III/Ghoul
- Henry Rollins as Benjamin Knox/Bonk
- Rachael Leigh Cook as Chelsea Cunningham
- Lauren Tom as Dana Tan
- Mary Scheer as Mrs. Drake
- Vernee Watson-Johnson as Ms. Joyce Carr