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- An unnamed Warrior is the playable Xenomorph character in the game. A both intelligent and aggressive Xenomorph, it caused the incident on LV-1201 in attempting to escape from the Primary Operations Complex back to the Hive.
- Corporal Andrew "Frosty" Harrison is the playable human character in the game. A Colonial Marine, he is characterized by a refusal to abandon his fellow Marines. Other than combat, Harrison specializes in electronics and is also qualified to pilot an elite "Exo-Suit".
- Prince is the playable Predator character in the game. 19 years before, he and his clan attacked a Weyland-Yutani outpost, where he chased and downed a Colonial Marine known as Rykov. Now, knowing Rykov is still alive, the Predator has come to LV-1201 to finish the job.
- Dr. Arnaud Eisenberg is the head researcher in charge of the LV-1201 facility. He was the only survivor of a disastrous first expedition; when a massive storm cut off his outpost's security systems and communications, he was forced to kill and eat his colleagues. He is the main antagonist of the alien campaign. He is trapped in the hive by the aliens.
- General Vassili Rykov is the head of the Iron Bears or "Corporates", a group of private military contractors employed by the Weyland-Yutani Corporation. He is the main antagonist of the Predator campaign. Killed by the Prince, who takes his spine as a trophy.
- The Empress is the primary Xenomorph Queen on LV-1201. Eisenberg's efforts are focussed on capturing and controlling her. She is larger than the younger Queens. She is the main antagonist of the Marine campaign.
- Major McCain is the commanding officer of the Colonial Marines dispatched to LV-1201 at Weyland-Yutani's request. He is a fine leader, but is oblivious to what Rykov and Eisenberg have in store for him and his Marines. He survives the mission.
- Duke Morrison is a Marine (seemingly of Latino descent despite his name) under the command of Major McCain. Both sarcastic and relatively cowardly, he is considered an annoyance by the other Marines (easily comparable to Private Hudson from Aliens). He survives the mission.
- Johnson is the premier dropship pilot aboard the USS Verloc. She has a strong British accent and a strong-willed personality, as shown in her response to Major McCain's question, "Where the hell did you learn to fly?"; "Flight school! The same place that taught me not to fly blind in hurricane winds!" She survives the mission with Duke, Blackwell, McCain, and Harrison
- Decker is a Marine private in Sergeant Hall's team. He is badly burnt in a Xenomorph ambush and is put in an APC to heal. He heals quickly, however, and is later seen guarding McCain and the APCs. He is killed by a Xenomorph Queen as Harrison enters the Hive. He resembles the Grenadier Marine character.
- Jones is an African-American Marine under the command of Major McCain. A heavy weapons specialist. He survives the mission.
- Sergeant Hall is a female African-American Marine. She is second-in-command to Major McCain. During the raid on the POC, her squad is ambushed and she is captured and impregnated by the Xenomorphs. She dies when it bursts through her ribcage.
- Sergeant Blackwell is a Marine under the command of Major McCain. He is head of Harrison's squad until they were separated. He and Shugi retreat back to McCain and guard him for the rest of the game. He survives the mission.
- Shugi is a Marine under the command of Major McCain. He is part of Harrison's squad until they were separated. He and Blackwell retreat to Major McCain and stay in a marine APC or Dropship for the majority of the game.
- Tomiko is said to be an assemblyman's daughter who signed up for an internship on LV-1201 to find out what had happened to her brother, Kenji, who she suspects was murdered. She blackmails Harrison into finding a disk with evidence on it and later helps him out.
- Captain Jonas Richter is the captain of the cargo vessel Aurora and often ferries questionable objects (such as Alien eggs) with no questions asked for Weyland Yutani, though he often scans them to get an idea of what Weyland Yutani is doing.
- Sergeant Cisco is a temperamental guard Sergeant. He gets angry at soldiers easily and has no friends in the Pods. He is killed when the player controlled alien bursts out of his chest.
- Lloyd is the security guard who escorts Richter to Eisenberg.
- Stiles is a technician who assists Harrison while he is in the Pods (before Dunya betrays him). He is easily recognized because of his goatee.
- Major Dunya is a Russian member of the Iron Bears. When Corporal Harrison disables the security network in the tunnels connecting the POC to the Pods, he inadvertently disabled the security at the Pods themselves as well. Dunya's fiancé Dimitri was killed and as a result, Dunya blamed Harrison for his death. She is the playable human character in Aliens versus Predator 2: Primal Hunt.
- Ivan Hushvatov is General Rykov's right hand man, who sees to it that all of the general's orders are followed. He is killed by Harrison later on in the Marine Campaign.
- Chekov is one of General Rykov's henchmen. While returning from escorting his superior to the Hive, he is killed by The Alien.
- Pamela is a doctor working in the pods. She is found by a rogue Predalien.
- Dimitri is a Russian foot soldier in the Iron Bears and is Dunya's fiancé. He is killed when out guarding a scientist outside the pod when Harrison shuts off the security grid.
- Joss Blaney is a POC guard who is seen killed in the Marine campaign.
- Kenji is Tomiko's brother. He is only mentioned in the Marine campaign.
- Light Predator #1 is a Predator who hunts humans in the POC. He was killed by the player Alien character in which he self-destructs.
- Light Predator #2 is one of the player's Predator's clan mates. He was captured by the corporates where he was taken for autopsy.
- Heavy Predator #1 is one of the player's Predator's clan mates. He was captured by the corporates where he was taken for autopsy.
- Light Predator #3 is a Light Predator who was killed by the player Alien character in the last Alien level. He tries to kill the Empress.
- Heavy Predator #2 is a Heavy Predator who was killed by the player Alien character in the last Alien level. He tries to kill the Empress.
- POC Predator is a Predator who appears in the first and sixth Marine level. He was killed by Harrison.
- Commander Liam Hadden is a POC guard-officer who appears in the second and third Alien level. He and other 4 guards monitors the POC.
- Ichiro is a Marine sniper. He appears in the multiplayer mode. An unused sound file implied that Ichiro was originally going to be featured in the campaign, but was scrapped by unclear reasons.
- Luis is a Marine technician.
- Carter is a Marine, who is killed by an Alien. He appears in the second Marine level.
- Mulder is a Marine, who is dragged by an Alien. He resembles Duke. He appears in the second Marine level.
- Quaid is a Marine, who is killed as he repairs a station. He resembles Harrison. He appears in the second Marine level.
- Dr. Adam Post is one of the scientists at the research facility. Although he is only mentioned in text logs in the game, he appears in person in the Primal Hunt expansion pack.
- Dr. Betheny Clements is a scientist at the research facility. According to text logs in the game, she had voiced reservations about Rykov and later fell victim when a Facehugger escaped under suspicious circumstances.
- Dr. Maurita is a scientist at the research facility. According to text logs in the game, he was initially reported as being transferred, but later revealed to have died. His death caused quite some rumors at the facility.
- McCullough was head of security during the first expedition to LV-1201, in 2221. According to text logs found in the game,he died when Xenomorphs attacked the party.
- Weller was a member of the first expedition to LV-1201 in 2221. He was killed by Runners, according to text logs in the game.