Polterkitty is first seen in a cutscene after Luigi rescues Toad, sitting on Hellen Gravely's lap while she watches Luigi's progress. After Luigi retrieves the 9F elevator button from Morty in Paranormal Productions, Polterkitty steals and eats the button as soon as it is placed in the elevator. Luigi must use his Dark-Light Device to reveal Polterkitty's invisible footprints which lead to her location. After Luigi flashes Polterkitty with his Strobulb, she will transform into a monstrous panther-like form and seal off the room. Luigi must face away from Polterkitty's perch to make her drop down for a sneak attack; if he turns around too soon, she'll immediately return to her perch and wait for him to turn his back again.
To defeat Polterkitty, Luigi or Gooigi needs to flash the Strobulb at her just before she attacks, which she indicated by her rearing up and spreading her forelimbs wide as her eyes flash pink. This will stun her long enough for Luigi or Gooigi to catch her with the Poltergust G-00. Polterkitty does not take damage from the suction; instead, she must be repeatedly slammed into the ground. When her HP is reduced to 1, Polterkitty loses one of her tails to the Poltergust G-00, allowing her to flee to the Garden Suites. Luigi must follow her and repeat the process of flushing her out and provoking her attacks to remove her other two tails; when she loses her last tail, Polterkitty spits up the 9F elevator button and retreats.
Polterkitty reappears after Luigi retrieves the 14F elevator button from Johnny Deepend in the Fitness Center, once again stealing the button after it is placed in the elevator. This time, Polterpup helps Luigi track down Polterkitty, and together they hunt her down in the same way as before. Polterkitty has regrown the tails she had lost previously and increased her HP to 100. She flees to The Spectral Catch after losing one tail, then moves to the Twisted Suites when she loses the second. When Polterkitty is cornered for the third and final time, she attempts a fake-out of her own; as soon as Luigi turns to counter her attack, she quickly goes right behind him to attempt another slash attack. If Luigi reacts to this in time to stun Polterkitty, he will have a chance to completely suck her into the Poltergust G-00, retrieving the stolen elevator button and ending her mischief for good.
Upon confronting Hellen Gravely in the Master Suite, she specifically mentions Polterkitty's capture as one of her many grievances against Luigi prior to their battle.
In the Gallery, Polterkitty shares the same containment jar as Hellen Gravely, although she will not appear until the latter is captured.