There are unused audio files in Portal 2 that suggest there was once a scene where Caroline was forcibly transformed into GLaDOS.

'No listen to me! Sir, I do not want this.'
'I do not want this!'
'Mr. Johnson, I don't want this!'
'Cave, I'm begging you, don't! Please!'
'NOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOO!'
*GLaDOS voice* 'I am awake. And I am going to kill you all.'

From 2014 to 2016, The Game Awards featured a "Best Fan Creation" category. The 2014 winner was "Twitch Plays Pokémon" (a social experiment featuring a crowdsourced attempt to play through various Pokémon games), and the 2015 winner was "Portal Stories: Mel" (a modification of Portal 2). During 2016, however, two of the nominees for the category were removed prior to the show's airing: AM2R (an unofficial remake of Metroid II: Return of Samus) and Pokémon Uranium (a fan-made video game based on the Pokémon series). During a stream prior to the event, host Geoff Keighley explained that this was due to the fangames not being legally cleared by Nintendo, who owned the rights to the games they were based on. Additionally, "Best Fan Creation" would end up being the only category to not have its winner (Enderal: The Shards of Order, a conversion mod for The Elder Scrolls V: Skyrim) announced during the ceremony itself.

There are unused lines for Cave Johnson found within the games '.nut' files. The lines would've been used for a scene where the player finds an AI cube with Cave Johnson's conscious programmed inside it. He would meet GLaDOS and reveal that she indeed used to be Caroline and that he "volunteered" her to be a test subject to see if his conscious could be loaded into a machine. He would then continuously ask the player to kill him.

There was originally going to be a song featured in the game called "GLaDOS' Song". It was written and performed by Ellen McLain, the voice actor for GLaDOS. She wrote the song for use as an Easter Egg in the game but it was ultimately never used.

The song was actually recorded after the game's release, and was performed by both Ellen and her husband, John Lowrie, the voice of the Sniper from Team Fortress 2. It was later modified by Harry Callaghan to sound closer to GLaDOS' synthesized voice.

After the Authoring Tools update, the Adhesion Gel (which allowed Chell to defy gravity) was replaced with the Reflection Gel, a gel that reflects lasers. It also reflects the turrets' laser sights, but this is just a cosmetic feature and doesn't affect their actual "vision".

There is an unused Turret hidden in the game files called the Hover Turret. It has no programming other than to follow the player with its laser sight. It also has no model, so it shows up as a default ERROR model.

In an earlier version of the co-op plot, GLaDOS sends Atlas and P-Body out to find "artifacts" of the long-dead Aperture workers (coffee cups and such) to make them more human. Unused audio files from this period of development can still be found within the games files, revealing that in one of the rooms a comic strip is found, presumably Garfield from how it is described.

During the development of Portal 2, playtesters found GLaDOS to be far too mean and vindictive. The developers concluded that a shift in personality was necessary, to better reflect the change in the power dynamic between Chell and GLaDOS. As a result, some of her dialogue was changed to make GLaDOS more amicable and passive-aggressive.

When Portal 2's audio files are tampered with or deleted, in the place of a broken audio clip, the game will instead play a file called "error.wav". This audio clip is of Dr. Kleiner from Half-Life 2, taken from a scene in which Dr. Kleiner is beginning to teleport Alyx but the cord gets unplugged, saying "Oh fiddlesticks, what now?"

During Chapter 2, it is possible to grab Wheatley as he comes out of the wall in Test Chambers 2 and 7. He cannot be harmed by lasers and will not be destroyed by the Emancipation Grill, but disappears after taking the elevator to the next Test Chamber.

The paintings in Chell's room at the beginning of the game changes from a day scene to a night scene depending on whether it is daytime or nighttime.

There is an achievement/trophy in the game called Ship Overboard. In the back of an office in Chapter 6: The Fall, Spoiler:there is a secret door hiding a hallway which leads to a dry dock containing a life preserver from The Borealis, a scientific research vessel owned by Aperture Science. The Borealis was first featured in Half Life 2 Episode 2. Getting close to and looking at the life preserver will get you the achievement.

The Turret Opera, the song that was played at the end of the game, was sung entirely by Ellen McLain, who voiced both GlaDOS and the turrets. When recording the song, Ellen was asked to make up some words to put in the song, which she did using her "very bad high school Italian."

In Chamber 7, there is some sort of robotic singing coming from the Companion Cubes.

When the Companion Cube is destroyed, you can hear human screaming as it disintegrates. This, and GLaDOS' insistence that the cubes do not talk in the first game, as well as Doug Rattman being able to hear the cube speak, suggest that the cubes contain people.

In the final Doug Rattmann den in Portal 2, which is accessible through chapter 3 in test 17, if you go near the graffiti art you can hear somebody rambling through the wall, which is believed to be Doug himself.

In test chamber 8, GLaDOS will leave shortly after you start the test to fix the turbines. Before leaving, she will give you sped up instructions. If you slow down what she says, you will hear she is actually quoting a line from Moby Dick.

Pneumatic Diversity Vent was introduced in a trailer at E3 as a way to move stuff around like turrets. However nowhere in the game can it be found. It was cut because the puzzle aspect was very limiting.

According to Portal 2 writer Erik Wolpaw, Valve originally planned to include a gel that let you walk on walls.

Unfortunately, players would become queasy with its use. Valve tried reducing the frame rate and camera movement, but were eventually forced to drop the feature entirely.

The original idea for the game was to replace Chell with a new protagonist named Mel. Cave Johnson was to replace GLaDOS as the antagonist, and Mel was going to have many different personality spheres follow her (such as the Adventure Sphere, Betty, and one referred to as the "Morgan Freeman Sphere") instead of just Wheatley. Play testers were up for the idea of Mel until they became disappointed when GLaDOS woke up and didn't recognize the player as the character that defeated her in the first game. Mel was then put in the multiplayer before being scrapped entirely and replaced with Atlas and P-Body.