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Saya Otonashi, the central character, is a seemingly ordinary teenage girl adopted and living with amnesia. As her memories return, her life is turned upside down. She was born in 1833, survives on the blood of others, and is destined to destroy chiropterans and her twin sister Diva. Aiding her in her quest are her adopted brothers Kai and Riku and her ever-faithful chevalier Hagi. She also is aided by David, her protector from the powerful organization Red Shield.

Diva also has her own helpers, starting with her five powerful chevaliers: Amshel, Solomon, Karl, James and Nathan. They also have their own human allies as part of the Cinq Flèches Group that they operate, including Van Argiano who helps them develop the Delta 67 substance that Cinq Flèches uses to create chiropteran.

Working against both sides are the Schiff, a group of escaped artificial chiropteran created by Diva's group. The Schiff attack both groups, believing that the blood of Saya, Diva, or one of their chevaliers will save them from a fatal and painful disease they call the Thorn.

Chiropterans are hematophagous bat-like creatures that live by feeding on human blood. They generally possess extraordinary speed, strength, and supernatural healing abilities. They are also capable of contorting and transforming various body parts in order to fly or have longer limbs and higher level chiropterans have the ability to disguise themselves as ordinary humans.

As chiropterans can heal almost instantly from most wounds, bullets are non-lethal and only useful in slowing them down. Even explosions do not seem to be capable of killing them, though if buried or sealed inside stone they will go into hibernation until freed.

Chiropterans are created from the blood of a queen and, ironically, it is also the primary method of killing them. When the blood of another queen is introduced into a chiropteran's bloodstream, it starts a crystallizing chain-reaction that destroys them. Thus, chiropterans created from Diva's blood can be killed by Saya's blood, and vice versa.

Through human experimentation with Diva's blood, a substance called "Delta 67" was created. When "Delta 67" is introduced into the blood stream of humans, they change into chiropterans. As they are all formed from Diva's blood, they can be killed through the induction of Saya's blood.

The most basic form of chiropteran that are created from the straight infusion of Delta 67 into a human host. The human host quickly loses their human intelligence and memories, transforming into a creature resembling a horse-headed gorilla with fangs. Phase 1 chiropteran live and act purely on instinct, hunting and killing humans, and sometimes animals, for their blood.

Through the research efforts of Dr. Boris and Amshel, higher level, artificial chiropteran were created from the Delta 67. These are collectively called the Schiff. Unlike Phase 1 chiropteran, the Schiff have human level intelligence and emotions, though somewhat stunted by having lived most of their lives caged. They are aggressive due to their training and subjection to experiments. Exposed to other ways of living after escaping the lab, they learn to adapt. The Schiff are unique among the chiropteran as they die when exposed to direct sunlight. They also suffer from a condition called Thorn, which slowly crystallizes their blood and kills them.

Diva's group uses the genetic template of Moses, the leader of the Schiff, to create an improved and obedient version of the Schiff called the Corpse Corps. Like the Schiff, the Corpse Corps are capable fighters with bladed weapons. They are marketed to various national military organizations as "chiropteran exterminators", despite being chiropteran themselves.

Chevaliers are the second highest level of chiropterans in terms of strength and ability. They are created when a human drinks the blood of a queen. A powerful bond exists between each chevalier and the Queen that sired them, compelling the chevaliers to serve and protect her at all costs.

Chevaliers have even greater speed, strength and endurance than regular chiropterans. Though they retain the appearance in body form and age that they had when they were transformed, chevaliers have vast shape-shifting abilities. They can transform into unique forms resembling regular chiropterans, either entirely or limited to specific body parts, and can transform into the humans they have fed upon. They heal instantly from almost any non-lethal wound, though severe wounds will temporarily cause an increase in their need for blood. As with regular chiropterans, chevaliers can be killed with the blood of the opposite queen; however, the crystallization takes longer and it is possible to prevent death by removing the crystallizing body parts. A chevalier can also be killed by beheading or burning their whole body.

Queens, also called the originals, are the first chiropterans. In any given generation, there are always two Queens, who are female twins born from one of the last Queens, and they are naturally antagonistic with one another. Saya and Diva are the current Queens of their species. They were discovered by Joel Goldschmidt in the 19th century inside a mysterious creature found in Iceland known as SAYA. In his experiment, he raised Saya like a daughter, but kept Diva locked away in a tower with virtually no human contact, or even a name.

All other chiropterans are born of the blood of Queens, either through the queen's own efforts or human experimentation. Unlike all other chiropterans, Queens appear to retain a human appearance at all times, though they can change their human appearances somewhat. Their blood can also crystallize and destroy any chiropteran of any level that was created by their sister queen.

Queens spend only a few years awake at any given time, between decades of hibernation within a cocoon. Queens have enhanced speed, strength, and endurance that surpasses those of their chevaliers and other chiropterans. They may also have some shape-shifting abilities, though during the series their abilities seem to be limited to modifying to their outward human appearance. The strength of a queen's traits are dependent on their feeding regularly on human blood. As such, Diva, who freely feeds on humans, can also freely use all of her abilities, while Saya, who avoids feeding, is usually only slightly above human level strength.

When a Queen becomes pregnant with the next generation of Queens, her blood will lose its dangerous qualities. A queen can only be impregnated by a chevalier created by her sister queen, ensuring the purity of the species is maintained.

Produced by Production I.G and Aniplex and directed by Junichi Fujisaku, Blood+ was inspired by the 2000 anime film Blood: The Last Vampire; however, there are only a few allusions and basic elements from the film. Fujisaku has been involved with both works, including acting as the director for Blood+ and writing the novelization of Blood: The Last Vampire.