The player is given a playing field, seen in the Nintendo DSi's bottom screen,[6] populated with viruses of three colors: red, blue, and yellow.

The player manipulates these capsules as they fall, moving them left or right and rotating them such that they become aligned alongside viruses of matching colors.

Critics generally praised the gameplay but lamented the lack of the multiplayer mode offered in earlier Dr. Mario games.

"[8] Daemon Hatfield of IGN scored the game 7.5 out of 10, stating the single-player experience is "solid" but "the lack of multiplayer hinders its lifespan.

Reviewer Jon Jordan called Dr. Mario Express "a great puzzle game," but criticized the presentation, particularly the small size of the playing field and the graphics, which he felt were "terribly old fashioned and badly animated."