The game builds upon the "Adventures of Captain Toad" levels in Super Mario 3D World, where the basic gameplay model had first been introduced. In the game, players control Captain Toad, a Toad, and his companion Toadette, attempting to safely navigate through various obstacles and reach a gold star at the end of each level.[3] Completing the original version allows players to see the link between Captain Toad: Treasure Tracker and Super Mario 3D World, and having save data from the latter or beating Treasure Tracker allows players to play four 3D World levels as Captain Toad. Meanwhile, the Switch version has an updated ending linked to Super Mario Odyssey and scanning the Wedding Amiibo unlocks four bonus levels adapted from Odyssey.

The player can manipulate the camera to view levels and potentially reveal hidden areas containing bonus items. Both characters can be hit once before dying, but collecting mushrooms restores their health. As in Super Mario 3D World, Captain Toad can also walk and run, but cannot jump due to his heavy backpack. As in Super Mario Bros. 2, he can pull up plants from the ground, granting the player coins or turnips to throw at enemies. He can also use the Super Pickax to clear out enemies and obstacle blocks, similar to the hammer in Donkey Kong. The game makes use of the Wii U GamePad's features, as the player can touch the screen to manipulate platforms, blow on the microphone to activate certain platforms, and use gyroscopic controls to aim turnips while riding mine carts.[4] The game makes use of amiibo: using the Toad amiibo places a "pixel Toad" in levels for Captain Toad to find; other figurines give the player extra lives