Bloodlines is a role-playing game with the choice between first person and third-person perspectives. The player character's ability to overcome obstacles is in many cases a mixture of player and character abilities, with character stats determining the effectiveness of actions, and player abilities determining whether or not the actions succeed. For example, the ability to move silently and avoid being detected is heavily influenced by the character's Dexterity and Stealth ratings; however, if the player does not stay in the shadows while sneaking past enemies, the character can still be detected.

The player character increases in power dramatically during the course of the game through the expenditure of earned experience points on attributes, skills, and vampire abilities called "Disciplines". A multitude of items, weapons, and books can be found or purchased to make the player character even more powerful. Melee and ranged weapons exist in equal numbers, although only in the later stages of the game.

How the player interacts with the game world varies depending upon which clan the player character belongs to. Differences range from different dialogue options becoming available to certain quests becoming available or unavailable. The most notable gameplay differences are experienced by those who play as Malkavian (due to their insanity, dialogue options are often non sequiturs, making it difficult to conduct conversations and negotiations; Malkavians also encounter numerous bizarre moments during gameplay, such as television sets and stop signs speaking to them) and Nosferatu (who, in order to avoid Masquerade violations, are prohibited from speaking to humans and who do not have access to any gameplay options involving seduction).

Unlike most role-playing video games, the experience needed to increase stats and skills is not awarded for killing enemies. Experience points are awarded solely for completing quests, no matter how many creatures the player eliminates in the process (though the quest objective often involves killing). This encourages the player to complete quests in creative ways and significantly increases the game's replay value.

The game invokes two other unique penalties and rewards for certain behaviors in the game's non-quest (i.e. non-combat) areas. Players are penalized for exhibiting vampiric abilities in front of humans by the loss of Masquerade points, which can also be reinstated by performing actions to protect the Masquerade. If the player loses 5 Masquerade points, the game ends. Also, the player is able to gain and lose "humanity" points, which have an impact on how well the character can be controlled when his or her blood supply is low. This can potentially cause the character to go into a feeding frenzy at the wrong time which in turn can lead to Masquerade violations. Humanity points are awarded for acts of kindness, such as finding alternatives to killing certain non-player characters. They can be taken away if the player character kills a human outside a combat zone (or even sometimes within a combat zone if the human is a noncombatant), intentionally or not, or if the character commits an unethical deed such as stealing money from a charity. Unlike Masquerade points, the game does not end if the player humanity level drops to minimum (3 by default), but the player's character is almost certain to enter frenzy when it is this low (and they are hungry, or take damage), and some dialog options may change. Experience points can be used to purchase humanity points. Having a very high or very low humanity affects some conversation options.