Holmes, 27, has pleaded not guilty by reason of insanity to 166 charges of murder, attempted murder and explosives offenses. He was arrested wearing a gas mask, helmet and body armor at the scene of the shooting rampage in the Denver suburb of Aurora. Seventy people were also hurt.
His lawyers have said that Holmes was suffering a "psychotic episode" at the time. If convicted, he could face the death penalty.
Jury selection may take up to four months as Arapahoe County District Court Judge Carlos Samour seeks to narrow the field to a panel of 12 jurors and 12 alternates.
The large number of people summoned reflects the difficulty expected in finding an impartial jury for such a high-profile, emotive case. Each person will be questioned about views on topics such as insanity defenses and the death penalty.
Holmes was in court for a pre-trial hearing on Tuesday, dressed for the first time in a gray suit jacket, open-neck blue shirt and khaki pants, as opposed to his usual red prison garb. Bearded and wearing spectacles, he chatted and smiled with his lawyers, while tethered to the floor below the desk.
Holmes, who first appeared in court in the days after the shooting looking dazed and with his hair dyed red-orange, is expected to be back in a courtroom later on Tuesday when the first group of 250 potential jurors will be addressed by the judge, watch a video on jury duty and fill out questionnaires.
The trial has been delayed several times, mostly by wrangles over Holmes' state of mind when he opened fire with a handgun, shotgun, and semi-automatic rifle inside the crowded premier of "The Dark Knight Rises" film at Aurora's Century 16 multiplex.
The Southern California native has undergone two-court ordered sanity exams since his arrest. The exams have produced dozens of hours of video and thousands of documents, all sealed by the judge; according to court papers they provided conflicting results.
HORRIFIC SCENES
At a preliminary hearing in January 2013, prosecutors gave a preview of the evidence that will be used against Holmes at trial.
First responders testified to horrific scenes in the body-strewn theater 9, where blood pooled on the floor while the movie still played, its soundtrack blaring. A fire alarm rang, strobe lights flashed, and wounded victims screamed.
One policeman said he initially mistook the gunman for a fellow officer because he was wearing body armor. But he soon realized it was not police-issue gear and saw Holmes was acting oddly. Holmes was ordered to the ground and handcuffed.
Another officer testified that when Holmes was then asked whether he had an accomplice, he replied: "No, it's just me."
Holmes has spoken just twice in open court, uttering one-word responses when questioned by the judge.
Holmes, who graduated with honors from the University of California, Riverside, has no criminal record and had been courted by the neuroscience doctoral programs of several universities. Shortly before the attack, he had withdrawn from a graduate program at the University of Colorado's Anschutz Medical Campus.
Holmes has been hospitalized at least twice since his arrest, according to his lawyers. Once he was treated for an apparent self-inflicted head injury. On another occasion he was taken to a Denver psychiatric ward where his lawyers said he was held "frequently in restraints" for several days.
Holmes also faces charges for allegedly rigging his apartment near the theater with explosives, which were defused by bomb technicians.
Colorado prosecutors rarely seek the death penalty, and the state has just three inmates on death row. Several death sentences have been commuted to life without parole thanks in part to the public defenders office, which is defending Holmes.
David Lane, a veteran Denver criminal defense lawyer who has tried some two dozen capital cases in several states and in federal court, said Colorado's public defenders have long been among the country's strongest death penalty defense attorneys.
"Their appellate office is second to none," Lane told Reuters. "Holmes has the best legal talent money can't buy, and he's extremely lucky."
