The player controls the movements of the eponymous Professor Layton and his young assistant Luke through several locations, unlike in the previous game which is confined to just one town.

Hamster toys are collected to help Luke give a morbidly obese hamster a workout; pieces of a shattered camera that Sammy accidentally dropped can be assembled to repair it; and players can earn tea ingredients to brew new recipes and serve cups of tea to Luke, Layton and people they meet.

They are followed by Inspector Chelmey, tracking down the crime, and Flora, who sneaks aboard the train but is eventually discovered by the pair.

As they enjoy the celebration, Layton and Luke learn that the town's founder, Sophia, also had an interest in the Elysian Box, but she died the year before, and her granddaughter Katia continues to seek it out.

Layton and Luke explore the town and learn it was founded on top of rich mine deposits by Duke Herzen and his sons, Anton and Fredrich.

Some fifty years ago, upon discovery of a new vein of gold, strange incidents began to occur around town, and many of its citizens left.

Fredrich left with his part of the family fortune and founding the Molentary Express, changing his name to hide his identity.

On returning to the hotel, Layton and Luke find that the remainder of the train's contingent has arrived, and Chelmey has arrested a conductor named Sammy as a suspect in the theft of the box and Schrader's death.

At the castle, the surprisingly young Anton initially welcomes them as his guests, but when they start to ask about the Elysian Box, he becomes suspicious and at one point ties the pair up, though they are able to escape.

Unfortunately, Anton lashes out with his saber in rage and disbelief, cutting a chain holding the chandelier in place and causing the castle to collapse.

Luke opens the special compartment and reveals that Sophia had gotten the box and left her own note to Anton, stating her love for him and Katia's relationship to her.

As Layton and his friends return to London, they learn that Dr. Schrader had only fallen into a temporary coma from his exposure to the gas from the box, and has now fully recovered.

[5] By this time, Level-5 had already decided upon the Japanese names of Curious Village and Professor Layton and the Unwound Future, but were originally planning to entitle the second game "Layton-kyōju to Yū-rei Jima no Himitsu".

[5] Level-5 also tried to update existing systems within the game, such as the Professor's suitcase and minigames; ultimately, Diabolical Box used up nearly twice as much data than its predecessor.

The international release of the game utilizes an instrumental version, though it similarly was not included on the album either due to not have being created at the time.

[33] At an interview following the conference, he confirmed that the name was the official English title and that the localization was currently being worked on, which he hoped to be finished in about six months.

"[10] Wired gave it a score of eight out of ten and said, "While Diabolical Box's gameplay, animation and plot are quite a bit like its predecessor's, slight improvements make this installment of the Professor Layton saga even more enjoyable than the last.

"[30] However, The Daily Telegraph gave it a score of seven out of ten and said that it "still has more charm and character than most and—despite the hiccups—provides a challenging, fun and satisfying puzzle experience for players young, old—and everything in between.

"[9] VideoGamer.com gave the game a 8/10, stating: "If you liked Mysterious Village and need more, or if you just want something genuinely fresh and original for your DS, Pandora's Box is the answer you're looking for.