The Binding of Isaac: Rebirth is a game based on the biblical story of the binding of Isaac. The story takes place in the ancient Middle East, about 400 years before the birth of Jesus. The main characters are the mother of Isaac, and the two sons, Abraham and Isaac. In the story, Abraham and Isaac are sent to the cave where the angel is waiting to take them away from this world to the next. But the angel's plan is interrupted by the arrival of a third character, the father of the twins, who is afraid that his two sons will die in the cave. So the father tries to persuade the twins to leave the cave with him. But when the twins refuse to leave, the father gives his son a new task to perform: he must find a ram to be placed in the breach where the twins will be. This ram is to be used as a substitute for the twins, who are to be saved from the wrath of God.

The plot of the game is based on the biblical story. The gameplay is designed to be a shooter.