In a desolate and empty New York City, a military virologist named Robert Neville is possibly the last human survivor of a pandemic that has transformed the populace into aggressive, nocturnal mutants called Darkseekers. The virus, originally created to cure cancer, spread rapidly and proved incurable, leading to the collapse of society. Neville, immune to the virus, spends his days scavenging for supplies, trying to find a cure, and broadcasting messages in hopes of finding other survivors, while his nights are spent locked securely in his heavily fortified Washington Square Park home.

Besides his own survival, Neville's mission is driven by the guilt of having not been able to save his family or stop the spread of the virus. He conducts experiments on captured Darkseekers, seeking a way to reverse their condition with his own immune blood. Accompanied only by his German Shepherd, Sam, Neville's isolation is palpable as he deals with the psychological toll of being alone. He sets up mannequins in the streets as a way to inject a semblance of normalcy and companionship into his life, clinging to a routine that keeps him from despair. Meanwhile, the Darkseekers, lurking in the shadows, constantly remind him of the dangers of this new world.