The ending to the Alternate Theatrical Version varies from the original ending. Instead of blowing himself and the hemocytes up with a grenade, Neville discovers that the hemocytes actually came for the female he captured earlier in the movie. He relieves her of the cure and returns her to the hemocytes. Afterwards, the hemocytes leave and let Neville, Anna, and Ethan be. The three then are seen leaving New York heading to Vermont's safe zone with Anna broadcasting a message telling all other survivors that she is with Neville and is heading to the safe zone.
The alternate theatrical version contains several new scenes:
- After Neville captures the Alpha Female, he notices that she has a butterfly tattoo on the back of her shoulder.
- After Neville thanks Anna for stitching his leg wound, they go out to where Neville was trapped the previous day. Neville recognizes the equipment as his own, but can't fathom what they're doing there. Anna asks him if the Dark Seekers could've made the trap, but Neville is adamant that they can't think or feel. Anna still brings up the point that Neville could be wrong and the Dark Seekers made the trap for him.
- Neville, Anna, and Ethan go to the Metropolitan Museum of Art. Neville and Anna sit together and watch Ethan fall into the water where the fish are. Ethan playfully splashes around, and Neville throws some food into the water. The fishes move frantically to get the food while Ethan floats in the middle of them. He remarks that it's cold, which makes Neville realize that he needs to lower the temperature of the Dark Seekers in order for the cure to work. He tells Anna and Ethan that they have to go.
- Additional shots of the Dark Seekers making their way into Neville's home.
- When the Dark Seekers corner Neville, Anna, and Ethan behind the Plexiglas, the Alpha Male shoves others out of his way and tries to break through the door. Neville shouts for him to stop, but the Alpha Male doesn't listen to him. Eventually, the Alpha Male stops on his own and uses his hand to smear the shape of a butterfly on the door. Neville sees it, and then sees the butterfly tattoo again on the Alpha Female's shoulder. He realizes that he just wants his girl back. Neville puts his pistol on the floor and unhooks the Alpha Female from his equipment. Anna asks him what he's doing, and Neville says that he's listening. He asks Anna to open the door. She grabs his pistol off the floor and reluctantly opens the door. Neville wheels the Alpha Female out into the lab, where the Alpha Male stands behind him and then growls next to his head. Anna closes the door and stays behind with Ethan. Neville tells the Alpha Male that he needs to get a syringe and slowly walks towards a drawer. Dark Seekers surround Neville and are ready to attack him, but the Alpha Male keeps them at bay. Neville gets the syringe he needs and injects the Alpha Female in the arm, waking her up. The Alpha Female hisses in delight upon seeing the Alpha Male, who tenderly presses their foreheads together. Neville is shocked that they feel love. The Alpha Male picks up his girl, screeches at Neville, and then leaves with the other Dark Seekers. Neville drops to the floor and looks at the wall of pictures he has of the Dark Seekers he's killed over the years, realizing that they have all been capable of thinking and feeling. The next day, Neville drives past the lion family he came across near the beginning when he was hunting the deer. Neville, Anna, and Ethan drive out of the city on the Washington Bridge. The film ends with Anna broadcasting a message to other possible survivors. She says that she's traveling with Robert Neville and a boy named Ethan. She says that they have a cure for the virus and are heading towards Vermont. She urges anyone who's listening to have faith and continue to listen for their future broadcasts.