Seven years after the Buffalo Bill case, FBI agent Clarice Starling witnesses her career crumble around her. During a botched drug raid, Starling kills a meth dealer, who was carrying a baby.^{[clarification needed]} Fugitive serial killer Hannibal Lecter, who has been living in Florence, Italy, under an assumed name, sends her a letter of condolence and requests more information about her personal life. Desperate to catch Lecter, the FBI tasks Starling with apprehending him. She meets with Barney Matthews, former orderly of Baltimore State Hospital for the Criminally Insane. When Barney asks Starling if she ever feared Lecter visiting her, she replied that she did not, as "he said he wouldn't".

Meanwhile, Mason Verger, a wealthy, sadistic pedophile whom Lecter disfigured during a therapy session years before, plans to get revenge by feeding Lecter to wild boars, using Starling as bait. He is aided by corrupt Justice Department agent Paul Krendler, Starling's nemesis. Rinaldo Pazzi, a disgraced Italian detective, pursues Lecter in the interests of collecting Verger's bounty on him. However, Lecter disembowels and hangs Pazzi in reference to the lynchings of the Pazzi conspirators. After killing one of Verger's men, Lecter escapes to the United States, where he begins pursuing Starling.

The novel briefly touches upon Lecter's childhood, specifically the death of his younger sister, Mischa. The two were orphaned during World War II, and a group of deserters killed and ate Mischa, something that haunts Lecter.

Barney briefly works for Verger, meeting Verger's sister and bodyguard Margot, a lesbian bodybuilder whom Verger molested and raped as a child. Her father disinherited her after learning of her homosexuality. Margot, who is infertile, tells him that she works for her brother because she needs Mason's sperm to have a child with her partner, Judy, and inherit the Verger family fortune.

Verger's men capture Lecter, and Starling pursues them. When Starling catches up to Lecter, Verger's men shoot her with tranquilizers. Verger unleashes his wild boars, but they lose interest in Lecter when they smell no fear on him, and turn on Verger's men. In the confusion, Lecter carries the unconscious Starling to safety and escapes with her. At the same time, Margot obtains Mason's sperm by sodomizing him with a cattle prod and kills him by shoving his pet moray eel down his throat. Lecter, who had briefly treated Margot after her brother abused her, had urged her to blame the murder on him, so she leaves a strand of Lecter's hair at the scene.

Using a regimen of psychotropic drugs and behavioral therapy, Lecter attempts to brainwash Starling, hoping to make her believe she is Mischa, returned to life. She ultimately resists, however, and tells him that Mischa will have to live on within him. Lecter captures Krendler and lobotomizes him, and then he and Starling eat Krendler's prefrontal cortex before Lecter kills him. Starling undresses and offers her breasts to Lecter, which he accepts. The two then become lovers and disappear together.

Three years later, Barney and his girlfriend go to Buenos Aires to see a Vermeer painting. At the opera, Barney spots Lecter and Starling; fearing for his life, he flees with his girlfriend.