Spano was born in San Francisco, California, the son of Virginia Jean (née Carpenter) and Vincent Dante Spano, a physician. He graduated from Archbishop Riordan High School in 1963, and he is an honorary member of the House of Russi. Spano and his wife Joan Zerrien, a therapist, were married in 1980. They have two adopted daughters.

Joe Spano was a member of the San Francisco improv group The Wing, and in college debuted as Paris in a production of Romeo and Juliet in 1967. In 1968, he helped found the Berkeley Repertory Theatre, appearing in its first production, and stayed with the company for 10 years. He moved to Hollywood in the late 1970s, landing guest shots on TV and bit roles in American Graffiti (1973) and The Enforcer (1976/I).

He is a veteran stage actor on the east and west coasts. Spano made his Broadway debut in 1992 in the Roundabout Theater revival of Arthur Miller's The Price, with Eli Wallach, which was nominated for a Tony for Best Revival. West coast stage credits include Eduardo Pavlovsky's Potestad, and David Mamet's Speed-the-Plow and American Buffalo, for which he was awarded an LA Drama Critics Circle Award. At the Rubicon Theater in Ventura he has played General Burgoyne in Shaw's The Devil's Disciple, Greg in A. R. Gurney's Sylvia and Vladimir in Waiting for Godot. He is a member of the Antaeus Theater Company and a founding member of three other theater companies. He played a seductive vampire in the cult musical Dracula: A Musical Nightmare in a small LA theatre. He also appeared in the TV movie Brotherhood of Justice with Keanu Reeves and Kiefer Sutherland.

After Hill Street Blues ended, Spano won recurring roles in television police shows Murder One (1995) and NYPD Blue (1993), again as a detective, and has appeared regularly in 27 television movies and 20 television shows like The X-Files (episodes Tempus Fugit and Max), Mercy Point and Amazing Grace. Spano won the Emmy award in 1988 for Best Guest Actor in a Drama Series for a role he played in an episode of Midnight Caller. He has appeared in 27 feature films, including working alongside Tom Hanks in Apollo 13 and Richard Gere and Edward Norton in Primal Fear. His credits are often confused with Australian actor Joseph Spano. They are not related.