If it is either Halloween or a Full Moon, you can wear special items for each class. If you put the Grand Duchess costume set on the Heavy he will have a large amount of new dialogue to say, one of which is "Everyone! Friendship is stupid magic!" Referencing the cartoon My Little Pony: Friendship is Magic.

The in-game hat item "The Tavish DeGroot Experience" is a direct reference to Jimi Hendrix. It is named after his band, "The Jimi Hendrix Experience", and is a similar style to the hat that he was often seen wearing. The item's description is also a reference to Hendrix's first single, "Hey Joe", and reads "Hey, Joe, where you going with that sticky launcher in your hand?".

Early during development, Team Fortress 2's visual style appeared different than its final cartoon appearance. It was similar to its predecessor, Team Fortress Classic, and the Counter-Strike series.

The Buffalo Steak Sandvich was originally going to be a peyote cactus. Playtesters felt that a man in a native American headdress punching people in a peyote-induced rage would be considered racist. The creator had to quickly replace the item with something else.

The Engineer Update was originally going to have a new building called the Repair Node, which would repair nearby buildings in bursts. It was scrapped due to how poorly the Engineer played when it replaced the Dispenser or the Teleporter.

The Mann vs. Machine update, released in August of 2012, had been teased by Valve for almost 3 years:
• In October of 2009, during the Haunted Hallowe'en Special, the will of Zepheniah Mann, the father of Redmond and Blutarch Mann, the owners of Blu and Red, respectively, showed that he had an unknown third son, who was revealed during the Mann vs Machine update as Gray Mann, the one sending robots to destroy Mann Co.
• During the 119th update several months later, a teaser video was released for the Engineer Update. In said video, multiple robot heads of the Heavy and Demoman classes could be seen in the background. These would later be seen as the heads for their respective robots sent by Gray Mann.
• Much later, during the Christmas 2011 update, a hidden back room, accessible only via cheats, was found in the newly added map "Foundry". In the room, it shows a cowering Heavy and Scout looking at developed pictures showing several of Gray Mann's robots. Besides these pictures, there are also 3 numbers in the map, "13 22 13", which in alpha numeric code translates to "MVM", the initals of the future update.
• During the Pyromania update 6 months later, the also newly added map "Doomsday" included another room only accessible with cheats. In this room, a disappointed Spy can be seen with the same 4 pictures and a map showing a town, a factory, and a farm. These 3 places would be the 3 MVM maps, Coaltown, Mannworks, and Decoy, respectively.
• 2 months later, the actual Mann vs. Machine update was released

If a Scout is blown up, there is a 1 in 100 chance that a dove will fly out of his body. This is a reference to the Meet the Medic trailer, in which the medic accidentally seals his pet bird, Archimedes, inside the scout's chest.

On the official Team Fortress 2 Wiki, pressing up, up, down, down, left, right, left, right, B, A, enter will cause a Spy to drop from above, plant a sapper on the logo, and audio will play from the Engineer.

There is a piece of unused music that was originally intended to play in the background of the Upgrade Station. While cut from the final version, it is still available within the game's files.

The SMG was originally intended for the Scout. Not only does the first trailer show off the Scout running with the SMG, but the game files for the weapon are still named "tf_weapon_smg_scout".

In early development, Team Fortress 2 contained grenades. The commentary for the game states that they didn't use grenades in the end because they were afraid that players would just spam them, so they were removed from the game entirely.

The Red Tape Recorder when slowed down to -80% says, "Let's make some noise"

The Pain Train, a melee weapon used by the Soldier and the Demoman, was originally intended for the Pyro.

The bow tie "Dr. Whoa" is a reference to the science fiction series "Doctor Who" and is often associated with the main character 'the Doctor', particularly the 11th incarnation. The bow tie can be worn by either the Medic or Spy and was created through the Steam Workshop by the user "TauVee".

The Proof Of Purchase, a promotional hat, is a reference to the human player model from an early version of the game named "Invasion." The hat's description further supports this, stating "From another era."

The Spy knife "Your Eternal Reward" is a reference to a scene in the 1992 film Aladdin, in which Jafar fools Aladdin with his disguise and then attempts to murder him with a dagger that Jafar refers to as "Your Eternal Reward."

The Dalokohs Bar got its name from the Russian word for chocolate, spelled backwards (shokolad).

Most of the sounds for the Soldier's rocket launcher "The Original" are remastered from the original Quake rocket launcher.

As part of the Pyromania update, two new Scout weapons were the Baby Face Blaster and the Pretty Boy's Pocket Pistol. These are references to famous criminals from the 1930's. The criminals were nicknamed George "Baby Face" Nelson and Charles "Pretty Boy" Floyd.

The Medic's pet bird, Archimedes, is named after the famous Greek physicist of the same name.