These are what we call the 'YMMV items.' Things that some people find in this work. We call them 'your mileage might vary' because not everyone sees these things in the same way. This starts discussions in the trope lists, a thing we don't want. Please use the discussion page if you'd like to discuss any of these items.
Nightmare Fuel: Dimitri's flashback in which he discovers Claire's dead body. All you see is her arm, but it's still unsettling, especially when one remembers this is a game for children.
A mentally unbalanced Clive using a mobile fortress to devastate London. It's not made clear if the area was evacuated, and if it wasn't, he may have killed a large number of people, although it's not brought up in-game.
Subject 3's backstory. He was a cute little bunny, abandoned and orphaned, before he was captured and sold to scientists and subjected to a number of torturous experiments.
Tear Jerker: The ending. First, getting to know why Clive did all this - he lost his parents in a time machine accident. Then, the scene where Claire says goodbye to the professor. Note: Layton actually cries and takes off his hat during this scene. And finally... Luke leaving because his father got a new job, which is even more sad after you've played the 4th game and realize that Layton is also losing one of his best friends - Clark.
This exchange, when Luke and Layton are parting ways and Luke begins to cry
Layton: Now Luke, a gentleman does not make a scene in public.
Luke: I'm sorry, professor. I'm not a gentleman yet! *hugs him*
Dmitri's flashback to the day Claire died, after he arrived too late. God, he looks so broken during the entire scene.