Synopsis-In Brooklyn, Larry Daley's unstable work history makes his ex-wife, Erica, consider him a bad example to their ten-year-old son, Nick. Larry worries that Nick admires Erica's boyfriend Don more than him. To improve his financial stability, Larry applies for a job as night security guard at the Museum of Natural History. Retiring guard Cecil Fredericks hires Larry, despite his unpromising résumé. The museum, facing declining attendance and revenue, is replacing Cecil and his two colleagues, Gus and Reginald, with only one guard. Cecil gives Larry an instruction manual on museum security and warns him not to let anything "in... or out".
On his first night, Larry discovers that after sunset, the exhibits come to life, including a playful Tyrannosaurus skeleton nicknamed "Rexy"; a mischievous stuffed capuchin monkey named Dexter; various African animals; rival miniature civilizations portraying the Old West, Ancient Rome, and Ancient Maya; an Easter Island Moai obsessed with chewing gum; wax models of various historical figures, including the violent Attila the Hun; and four pyromaniacal Neanderthals. There is also Sacagawea, who, unlike the other exhibits, is in a glassed-enclosed diorama. A mounted Teddy Roosevelt rescues Larry from miniature leaders Jedediah and Octavius, who are attacking him. Teddy explains that ever since an ancient Egyptian artifact — the Golden Tablet of Pharaoh Ahkmenrah — was brought to the museum in 1952, the exhibits have come to life each night. If any exhibit figure is outside the museum at sunrise, they turn to dust. As Teddy helps Larry restore order, Larry learns that Teddy loves Sacagawea but is too shy to speak to her through the glass.
The next morning, Cecil, Reginald, and Gus return to check on Larry, but Larry says he is quitting. When Nick and Don stop by the museum to congratulate Larry on his new job, Larry decides to stay for his son's sake. To help him with the job, Cecil advises reading up on history.
That night, Larry better controls the exhibits, but the four Neanderthals set their display on fire while Dexter steals Larry's keys. As Larry extinguishes the fire, Dexter unlocks a window. One of the Neanderthals sees a trash bin fire on the street below and leaps out the window. Larry, frustrated with everything going awry, once again decides to quit. As he walks out, the Christopher Columbus statue gestures to where the Neanderthal escaped. Larry runs outside just as the rising sun disintegrates the Neanderthal into dust. Later that morning, museum director Dr. McPhee fires Larry over the damaged Neanderthal exhibit, though Larry convinces him to reconsider. Larry meets Rebecca Huntman, an attractive museum guide and historian. When he offers to introduce her to Sacagawea to help with her doctoral research, Rebecca believes Larry is mocking her as being a history geek.
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