Derek Bliss, a dedicated vampire hunter, begins his night on the streets by searching for a prostitute, only to confront a dangerous man threatening her with a razor blade. In a split second, Derek intervenes by pointing a uniquely shaped gun at his opponent's temple, causing the attacker to retreat. The prostitute, grateful for Derek's help, asks if there is anything she can do for him, but Derek swiftly demonstrates his lethal skills by shooting her with three stakes—revealing her as a vampire. Afterwards, he drags her body to an abandoned car lot and watches as the sunlight ignites her corpse into flames, capturing the grim scene on camera.
Later, Derek receives a call from the leader of the Van Helsing Group, who informs him of a new assignment. He travels to Mexico, where he visits a monastery housing a secretive group of vampire-hunting priests. During a strange vision, one of the priests suggests that the vampires might be attempting to link up with Derek himself, hinting at an ominous connection.
That night, the vampire princess Una and her fledglings launch a brutal attack on an innocent man, feeding on him in a fierce assault. The following day, Derek sets out to locate people from a list he has been given, but most of them are either dead or severely disturbed, with only a few remaining alive—who are quickly killed. At a coffee shop, Derek asks about a man named Jesse. While waiting, he converses with a woman named Zoey, who asks if Derek works in the "undead" business, instantly arousing his suspicion. Using a special lens, he confirms Zoey's nature as a vampire, lamenting her cuteness but quickly realizing her danger. Just as Derek is about to leave the bathroom, Una storms in, slashes the throats of all the customers, and kidnaps Jesse. When Derek emerges, he sees that Zoey has also fled, leading him to believe she was involved.
The next day, Derek spots Zoey on the side of the road during daylight hours. An argument ensues, where Zoey reveals she takes special pills to suppress her vampire urges. She admits she was bitten during a one-night stand, and she had a vision of the monastery previously visited by Derek. Both realize they are connected to a growing threat involving the legendary Béziers Cross—an artifact used in a ritual to grant vampires immunity from sunlight, making them virtually invincible, echoing the dark history of its use in the first film. They return to the monastery, only to find it slaughtered except for one surviving priest, Father Rodrigo. He shows Derek a makeshift vampire hunter's vehicle equipped with weapons and tools designed for their mission.
As they investigate, Derek encounters Sancho, a young boy with a permission slip from his mother, eager to join the hunt. The team is soon joined by Ray Collins from Memphis, who soon becomes entangled in the dangerous pursuit. During their mission, Una seduces Ray and manipulates him into leaving her pills where she can access them, further complicating their efforts. When the group reaches a village hiding vampires, they are cautiously welcomed, but Una succeeds in kidnapping Zoey after she regains her ability to walk in daylight. Derek, suspecting Sancho might have given her the pills, threatens him, but chaos erupts when a villager kills Ray after discovering the truth.
Derek and his team launch a desperate rescue mission for Zoey, ultimately extracting her from Una's clutches and returning her to a clinic. There, her vampire blood is exchanged for human blood as a temporary suppression method, since her pills have run out. Meanwhile, Derek devises a plan to save Father Rodrigo, who is actually not a true priest, revealing the dark deception. Una, unbothered by the pursuit, sets a fiery trap, but Derek manages to save Rodrigo from her flames.
The climax intensifies as Derek and his allies confront Una, nearly killing her in daylight, but she manages to escape. She captures Derek but is ultimately shot in the head with a shotgun by Derek, sending her head into the sun where it catches fire. Her body turns to stone with a black, beating heart visible in her chest, prompting Derek to deliver a final stake to her heart. In the aftermath, Sancho and Father Rodrigo decide to stay behind in the vampire-infested village, while Derek and Zoey drive into the sunset, ready to resupply on pills and continue their fight against the undead.
Throughout the film, the story explores themes of loyalty, sacrifice, and the eternal struggle between good and evil, portraying Derek Bliss as a relentless guardian in a world teetering between darkness and light.
