The story of Eternal Darkness takes place over four principal locations which the game skips back and forth between. They include an underground temple complex called the Forbidden City, in Persia; a Khmer temple in Angkor Thom, Cambodia; Oublié Cathedral in Amiens, France (not to be confused with Amiens Cathedral); and the Roivas Family Estate in Rhode Island, which also leads to an ancient underground city named Ehn'gha beneath the mansion. Each time a location is visited, it is done so in a different time period. Spanning from 26 BC to 2000 AD. Almost half of which take place in the 20th century. Each different era and character offers a different periodic and personal perspective on the location.

The chapters found in the game are not discovered in chronological order. Instead, to make the narrative more dramatic, each chapter jumps around the timeline of the plot. However, despite the overall story skipping back and forth through time, the chapters do follow chronological order within their respective locations. This is because each setting also has its own contained story.

In 2000 AD, Alexandra Roivas finds herself returning to her family's estate in Rhode Island after her grandfather, Edward Roivas, her only living relative, is found brutally murdered. Two weeks after returning, with the local police having gotten nowhere with the investigation, Alex decides to investigate the mansion for clues, and stumbles upon a secret room containing a book bound with human skin called the Tome of Eternal Darkness. Deciding to read it, she finds it contains accounts of various people in the past, beginning with the story of Pious Augustus.