Scully is a medical doctor and FBI agent, the partner of Special Agent Fox Mulder. She is Catholic, which often creates moral dilemmas for her, and is far more skeptical than Mulder, even in defiance of apparent evidence. She earned a B.S. in Physics from the University of Maryland, College Park. Her undergraduate thesis was entitled "Einstein's Twin Paradox: A New Interpretation."

Scully has an older brother, Bill, an older sister, Melissa, and a younger brother named Charles (or Charlie), who is never seen on the show. Her parents are Maggie and William, the latter of whom passed away of a massive coronary in the critically-acclaimed first season episode Beyond the Sea. She called her father "Ahab" from the book Moby-Dick. In return, he called her "Starbuck".

As with Mulder, work takes up a major part of her life, and she rarely dates or spends time with friends. In fact, she is given the nickname Mrs. Spooky early in the show for her position with Mulder. She and her partner are often mistaken for a couple.

Scully has a passing understanding of German (Unruhe), and wears a small crucifix necklace. She had a small tattoo of an Ouroboros snake on the small of her back (Never Again) but had it removed shortly afterwards (it cannot be seen when she is taken out of the Cryopod in the movie The X-Files: Fight The Future or when she is chained to a bed in Roadrunners, 8x05, though this could just be a continuity error). She owned a Pomeranian dog named Queequeg, after a character in Moby-Dick, which her father used to read to her. Queequeg met an untimely demise in Quagmire, eaten by either an alligator or a sea monster. She lives alone in a small apartment (Apt 35, 1419 of an unknown street, Georgetown, DC) near Washington, DC where a surprising number of people have died.

In Ascension, Scully is kidnapped by an ex-FBI mental patient named Duane Barry, and gets kidnapped by a military covert operation that were working with the alien conspirators, and is later returned (One Breath). She later finds out that a super hi-tech microchip was implanted in the back of her neck. After having it removed, she developed cancer in the fourth season (Memento Mori). She is hospitalized after her cancer becomes terminal (Redux). She is saved when Mulder breaks into the Department of Defense to retrieve another chip to be implanted back into her neck. (It should be mentioned that, at the time, Scully was also undergoing experimental medical treatments and was having a dramatic renewal of her faith.)

After being pronounced infertile, Scully became pregnant in the show's seventh season finale, Requiem. The child, named William after his grandfathers (Mulder is presumably the child's father), was born at the end of season 8. It was around this time that Mulder was fired from the FBI by Deputy Director Kersh, and Scully left the field to teach Forensics at Quantico. William was given up for adoption during the ninth season after Scully felt she could no longer provide the safety that William needed. William was a "miracle child", of some importance to the Alien Conspiracy. He demonstrated extraordinary powers, including telekinesis.