Chambers was born in Springfield, Ohio, the son of Pamela Sue (Willman) and John William Eugene Chambers II. He has a fraternal twin brother named Jason, one older brother John Jr. (Chip), and two older sisters, Mia and Susan. He and his twin brother visited hospitals often as children due to frequent bouts with pneumonia. He attended Southeastern High School in South Charleston, Ohio, and later Springfield Clark JVS.
In 1993, he married Keisha Chambers, a former model agency booker; they have five children together: Isabella (b. December 1994), twins Maya and Kaila (b. June 1997), Eva (b. March 1999) and Jackson (b. January 2002). In late January 2008, Chambers admitted that he suffered from a biological sleep disorder. The actor checked himself into Los Angeles's UCLA Medical Center to treat exhaustion due to the sleep disorder after a period of getting only two hours of sleep each week.