Tsukihime chronicles the story and life of a teenage boy named Shiki Tohno. As a young boy, Shiki suffered a life-threatening injury. As a result of this injury, Shiki became able to see "death lines" on objects and people; if Shiki traces these lines with a sharp object, the person or object that they belong to will be cleanly and easily cut. Soon after discovering this power, he is given a special pair of glasses by a mysterious woman. While wearing these glasses, Shiki cannot see the lines, and can thus live life as normal. Later, Shiki discovers that he can also see a place where the lines meet known as "points" on objects and people; these act as more immediate and powerful versions of the "lines." Eventually, it is revealed that Shiki is not merely killing a person or destroying an object when he cuts its lines or stabs its points, but is instead annihilating the victim/object's existence, therefore killing the victim/object as a side effect.

The game plot begins during Shiki's second year of high school. After spending eight years away from the Tohno household, which he was banished from after his injury, he moves back home following the death of the head of the Tohno family, Shiki's father. Shiki's sister, Akiha, assumes the responsibility of being the new head, and decides to let Shiki come back. As the story progresses, Shiki gradually discovers his mysterious past, and becomes implicated into strange adventures.

Tsukihime works as a normal visual novel. The player is presented with choices, some affect the story in large ways, others do not affect the story at all or affect it in small ways. Changes that affect Shiki's opinion on the characters and heroines usually change the direction of the story drastically. There are two scenarios: the Near-Side Route (as in "near-moon") which includes Arcueid and Ciel as selectable heroines, and the Far Side Route (as in far-moon) which includes Akiha, Hisui, and Kohaku as selectable heroines.

The protagonist of the series. He has Mystic Eyes of Death Perception, which allow him to see the death of things in the form of "lines" and "points" on objects and people; cutting the lines destroys the portion cut, while piercing the point destroys the existence of the object itself. He carries a pocketknife named Nanatsu-yoru (later learned that it can also be pronounced "Nanaya") and is quite skilled in its use. He has been living with the Arima family, one of the branching families of Tohno, since an accident 8 years ago. At the beginning of the game, he moves back home with his sister, Akiha.

Young Arcueid Brunestud and Zelretch

A beautiful blonde with ruby red eyes, Arcueid (or just "Arc" sometimes) is a mysterious vampire princess. But since she is a True Ancestor, she lacks some vampiric qualities, such as that of needing to consume blood to survive, and being nocturnal. She seems to be quite knowledgeable about many things, but is very naive when it comes to modern ideas. She is killed by Shiki in the first chapter, but she doesn't actually die (because she's a vampire) and, in turn, recruits Shiki to help her fight Roa. Arcueid was originally created as nothing more than a weapon to eliminate the True Ancestors that had fallen to their vampiric impulses. She was taught nothing except what was necessary to eliminate her target, and when she returned to the Millennium Castle even that knowledge would be erased before she was put to sleep. Originally did not have the vampiric impulses that all True Ancestors had, but this changed when Roa tricked her into drinking blood hidden within a rose, as she did not know she was not supposed to do that. After that, Roa, turning into a Dead Apostle, used his power gained by having his blood drunk by the strongest True Ancestor and wiped out most of the True Ancestors. From then on Arcueid uses a large portion of her power to suppress her vampiric impulses. She also changed her goal from killing fallen True Ancestors to killing Roa every time he reincarnates. By the time of Tsukihime, due to the weak state that Shiki left her in by "killing" her, she is only at 30% of her full power as she must use the remaining 70% to suppress her vampiric impulses. She is one of the two True Ancestors capable of using the Marble Phantasm, which allows the user to re-create the effect of anything that is naturally possible, (not counting the creation of living beings) and summon the Millennium Castle Brunestud, the home of the True Ancestors.

The sole member of the Japanese tea ceremony club and an upperclassman of Shiki's, or so it seems. She is actually the seventh most skilled executor for "The Burial Agency", a shadow branch of the Church created to exterminate "heretics". Her body was used in Roa's last incarnation for its superior magic circuit. After Roa-Ciel had killed her parents and taken over the whole village, Arcueid had come to kill Roa. Due to an unknown cause, Arcueid had supposedly killed Roa-Ciel causing Roa to reincarnate, but Ciel had survived. Due to this "error" by the universe, she cannot be killed until Roa dies. If she were to suffer any sort of wound, or even killed, time would reverse itself and heal her body to "fix its mistake" (the only exception to this would be if Shiki stabs her point, therefore destroying her existence). The Church took her in to examine her body to find possible ways to destroy Roa, but after no solution could be found, they attempted many times to kill her, only to find that she would come back to life every time. It was then decided to make an exception for her (out of fear/lack of ideas) to allow her to become an assassin for The Burial Agency. Her signature weapons are pages from the Bible that she turns into nail-like throwing swords using her mana. Her most powerful (known) weapon is an Exorcising Artifact from the Church known as the Seventh Holy Scripture. She has no personal grudge against Arcueid (other than the fact that she's a vampire) until they start competing for Shiki. Her Burial Agency outfit is not a uniform, but the result of one of her hobbies. She has a strange obsession for curry and will agree to almost anything for food that is curry-related.

Shiki's younger sister and the current head of the Tohno family. She is very prim and proper and carries a grace and nobility about her. As the story progresses, the viewer discovers that she has had mysterious powers since childhood. When her Tohno blood awakens, her hair turns a crimson shade and she can literally suck the heat out of her surroundings, which creates a burning effect on the object.

The younger of the two twin maids in the Tohno mansion, Hisui is a childhood friend of Shiki. She wears a western maid uniform and attends to Shiki when he comes back to the Tohno mansion. She acts cold and unfeeling, but it is only an act to hide her kinder nature for the sake of her sister. Her name, Hisui, is the Japanese word for jade, referring to her eye color. She is a Synchronizer, a person that can give someone life energy through exchange of body fluids.

The older of the two twin maids in the Tohno mansion, Kohaku, along with Hisui, is a childhood friend of Shiki. She wears a Japanese kimono and is always seen to be smiling and cheerful, being especially gifted with medicine. She hides her tragic past behind a cheerful face and Hisui's cold demeanor. Her name, Kohaku, is the Japanese word for amber, referring to her eye color. Like Hisui, Kohaku is a Synchronizer.

Nicknamed "The Serpent" for his reincarnation ability, Roa was a priest of the church that sought immortality and tricked Arcueid into drinking blood hidden within a rose, turning Roa into one of the most powerful Dark Apostles which allowed him to kill most of the True Ancestors, and to the point where he was strong enough to defeat Altrouge Brunestud when she tried to "teach him a lesson" as the rest of the Dead Apostles saw him as an upstart. His original body was finally defeated by a coalition comprised of Arcueid and the Burial Agency, and he has been re-incarnating ever since then. By the time of Tsukihime he has already re-incarnated 17 times and been destroyed by Arcueid each time as she seeks to gain her stolen power back, and perhaps to achieve some measure of revenge. Roa has two requirements for choosing the host of his next reincarnation, his family must have a high social standing so he would be born into luxury, and a body with a high potential. His last host was Ciel, who was chosen in haste and thus was born into a bakery's family but instead had a body with an extremely high physical and magical potential. Can be considered a friend or acquaintance of Nrvnqsr. He only shows up in the Near Side of the Moon scenarios.

Pronounced "Nero Kaos", this vampire is known as the tenth of the twenty-seven Ancestors of Dead Apostles, the highest in rank of the vampires in Tsukihime. Has 666 beasts on his body, from deer and wolves to mythological creatures like unicorns. The only conventional way to kill him is to destroy all 666 beasts at once or he will constantly regenerate, however this is practically impossible. Roa taught him a reality marble known as "The soil of genesis" which requires a destructive power that can destroy a continent. It functions as some kind of cage or restraining device. Can be considered a friend of Roa. Nrvnqsr Chaos is a name that was given to him by the Church. When he was still human, his name was Fabro Rowan.

A mysterious red-haired woman who always carries a suitcase. She met with Shiki eight years prior to the start of the series, and to this day, is the only person he will call "Teacher". She gave Shiki a pair of glasses (stolen from her sister T�ko), the Magangoroshi (Mystic Eye Killer), that blocks out the death lines and allow him to live a normal life. Current title holder of Beef-Eating contest in an unspecified steak restaurant in Japan. Her nickname is "Blue Gunner."

Shiki's classmate and best friend. His parents died when he was young, and he lives with his sister, Ichiko. In Kohaku's scenario he isn't seen after the first day in the game.

The patriarch of the Tohno Family. Dies just prior to the events of Tsukihime. He severely abused Kohaku when she was young.

Shiki's classmate, who has had a crush on him since middle school. In the Far Side of the Moon scenarios she's revealed to have been turned into a vampire. Shiki is forced to kill her. During development, when Satsuki had her own scenario, she was to to be capable of a rare, powerful type of sorcery, a Reality Marble in the name of Depletion Garden (Kokatsu Teien?), which literally dries up the mana of the environment. In Melty Blood, this skill returned as her last resort in battle and was adapted to the fighting game to actually deal direct damage.

Any creature which either physically requires blood to survive or has a great need for blood. Vampires are divided into two categories: "True Ancestors" (Shinso?) and "Dead Apostles" (Shito?).

Pure-blooded vampires born into the vampiric race. They have no physical need for blood, but the urge to drink blood (the same as their creator, Crimson Moon) rests within them. The majority of True Ancestors use a large amount of their powers to suppress their urge to drink blood. Those who cannot resist their urges and become insane, blood-sucking monsters are called Demon Lords. True Ancestors can create Dead Apostles by injecting their human victims with their blood.

Vampires who were once human. They could either have had their blood sucked by a True Ancestor or changed themselves into vampires through magical research. While Dead Apostles are immortal, they are not eternal like the True Ancestors. This is because their originally human bodies rejects the stronger DNA of the vampiric race, and as a result, a Dead Apostle's genetic structure deteriorates over time. In order to remedy this, they must continuously drink blood in order to take in additional genetic material. Dead Apostles can create the Dead by injecting their victims with their blood.

Mindless soldiers under the Dead Apostle's command. They are relatively weak, but still strong enough to take down a human with ease. The majority of the blood they drink is transferred to the Dead Apostle controlling them, making them ideal for concealing one's location while securing a safe stash of nutrients. There is a 1/100 chance of a person becoming the Dead, and a 1/1000 chance that that same person will retain their will. If a victim of a Dead Apostle retains their will, then they also become a vampire; however, they are still under the influence of their parent vampire like the Dead.

A vampiric capability which alters reality within a given range. A vampire whose Marble Phantasm is strong enough to summon the Millennium Castle Brunestud is hailed as vampiric royalty and given the last name of "Brunestud".

There is a large difference between magic and sorcery in Tsukihime. Magic refers to something which defies logic, but can still be achieved by anyone given time using arcane methods. Sorcery refers to the miracles that completely defy reality that can never be achieved just through time and effort alone.

The Burial Agency is the strongest of all heretic-hunting units within the Vatican. Known for their destructive capabilities and ruthlessness, they are the most feared amongst all internationally operating heretic hunters.

Any human with non-human blood within them. The non-human blood, in most cases, will result in improved physical capabilities and certain special abilities at the cost of the person's mental stability. The process through which a human within a demonic clan loses his/her sanity and becomes a mindless killing demon is known as Inversion Impulse.

Any human who is born with an arcane capability without non-human blood within them. Psychics range widely in capability, from being able to see a person's past to setting off ignitions at will.

An extremely rare ability, it enables the user to see and destroy lines and points of death on anything existent. Since it erases existence itself, the item does not necessarily have to be physically manifested.