Rock Band, by the original developers behind the Guitar Hero games, takes the concept a step further. Instead of solely playing the guitar along with songs, this game allows players to start a full virtual band with up to four players, with different sequences for each of the three supported peripherals: guitar (electric and bass), drum and microphone. Gameplay is based on hitting scrolling notes on the screen in time (scrolling vertically for drum and guitar), and getting the right pitch for the vocals (scrolling horizontally at the top of the screen, along with the lyrics).

The game offers a Solo Tour Mode for a single player or a Band World Tour (as well as Quick Play) for two to four players. By doing well players earn cash to customize the characters in the Rock Shop, with tattoos, face paint, clothing and logos. Customized characters are automatically locked to an instrument. Other customizations include movements on stage, instruments, and the general physique and appearance in the game. To succeed, players need to reach a set score through their combined performances. Multipliers for playing perfectly are tracked for the individual instruments and there can also be separate difficulty levels. The Star Power from the Guitar Hero series has been replaced by Overdrive, a meter filled by not making any mistakes in certain sections of a song. In band play using Overdrive also increases the band's score multiplier.

Players who fail during the song are locked out and the audio mix takes over. Other band members can revive the player by using their Overdrive. Failed players will however drag down the band's overall performance, so it becomes important to bring back band members. Next to the Overdrive sections, there are also Union Phrases (score bonus and extra Overdrive) and sometimes Big Rock Endings at the end of a song where another bonus can be gained.

In the Tour Mode players visit 41 venues in seventeen different cities. After the initial venues, others need a certain numbers of fans to be unlocked and certain continents only become available when tour buses or private jets are acquired. Not every venue comes with a set list of tracks. They consists of single tracks, multiple tracks, custom setlists or mystery setlists. Performing poorly can also have the band lose fans. Extra competition is included to attract managers, roadies and deals with record labels.

Next to the main modes there is also a versus mode for the individual instruments (Tug of War), Score Duel (get the most points) and a Tutorial mode. The PlayStation 3 and Xbox 360 versions feature multiplayer games with new songs through downloadable content.

The European version is not sold as a single package. Instead, there is an instruments package and the game is sold separately, for a much higher price than the original US release.