He was born in 1913 in Cedar Rapids, Iowa, to Joseph Ervin DeFore (1878–1942), a railroad engineer, and Albina Sylvia Nezerka (1883–1975).
DeFore's multiple film appearances include: The Male Animal (1942), The Human Comedy (uncredited, 1943), A Guy Named Joe(1943), Thirty Seconds over Tokyo (1944), The Affairs of Susan (1945), You Came Along (1945), Without Reservations (1946), It Happened on 5th Avenue (1947), Ramrod (1947), Romance on the High Seas (1948), My Friend Irma (1949), Too Late for Tears(1949), Dark City (1950), Southside 1-1000 (1950), She's Working Her Way Through College (1952), The Guy Who Came Back (1951),A Girl in Every Port (1952), Jumping Jacks (1952), Battle Hymn (1957), A Time to Love and a Time to Die (1958), and The Facts of Life (1960).
On May 6, 1953, DeFore was honored on the television series, This Is Your Life, a show that surprised the honoree on nationwide live television with accolades from coworkers, friends, and family. DeFore was led by Ozzie Nelson to believe that preparations were underway for a live promotional spot for The Adventures of Ozzie and Harriet on Hollywood Boulevard when suddenly he was surprised to hear Ralph Edwards exclaim: "Don DeFore - This is your life!" DeFore admirably managed to retain his composure while moving to the El Capitan Theatre, from which came the remainder of the broadcast, which included friends and family members from his native Cedar Rapids.
DeFore is less known as an actor in radio. He appeared on such radio programs as Old Gold Comedy Theater and Lux Radio Theater. From 1954-55, he served as President of the National Academy of Television Arts & Sciences. He was instrumental in arranging for the Emmy Awards to be broadcast on national television for the first time on March 7, 1955. That year, DeFore was nominated for Best Supporting Actor in a Regular Series for his work on The Adventures of Ozzie and Harriet.In time though, the role of Thorny was superseded by Lyle Talbot as Joe Randolph, and Mary Jane Croft as his wife, Clara.
In 1956, DeFore was cast as the Reverend C. E. "Stoney" Jackson in the episode entitled "The Comeback" of the religion anthology series, Crossroads. In the story line, the Reverend Jackson provides spiritual insight to assist Lou Brissie, a former professional baseball player wounded during World War II, to regain his strength so that he can return to the game. Brissie is played by Chuck Connors, himself a former professional baseball player, later cast as Lucas McCain on ABC's The Rifleman. Grant Withers played Coach Whitey Martin, and Edd Byrnes, Rhys Williams, and Robert Fuller were cast as former soldiers in this episode. X Brands played another baseball player.
From 1961 to 1965, DeFore was a co-star of the television series, Hazel as "Mr. B." (George Baxter), employer of the spirited, domineering housekeeper Hazel Burke, played by Shirley Booth and based on a character in The Saturday Evening Post. Whitney Blake played Mrs. Dorothy Baxter, and Bobby Buntrock portrayed the Baxters' son, Harold. The series ran on prime time until 1966, but DeFore was not seen in the final 1965–1966 season, when the show moved from NBC to CBS. Instead, Ray Fulmer appeared in the role of Steve Baxter, George's younger brother, and Lynn Borden played Barbara Baxter, Steve's wife.
From 1957 to 1962, with his younger brother, Verne DeFore (1918–2005), he operated Don DeFore's Silver Banjo Barbecue in Frontierland of Disneyland Park in Anaheim,California. The DeFores were the only sole proprietors ever permitted by Walt Disney and Disneyland to operate an independent business and restaurant inside the theme park.
In 1942, DeFore married Marion Holmes (born Marion Holm, January 21, 1918, Chicago). She was a singer with the Henry Busse Orchestra from 1935 to 1939, and later with Art Kassel and his "Castles in the Air" from 1939 until their marriage.
Judy Garland was the maid of honor at the DeFore wedding on February 14, 1942. DeFore and his wife were longtime residents of the Mandeville Canyon section of Brentwood, and DeFore, an active Republican, was the first "Honorary Mayor" of Brentwood. They had five children: Penny (born 1943), David (1945), Dawn (1948), Ron (1950), and Amy (1959).
Don DeFore died on December 22, 1993 at the age of eighty. He is interred at the Westwood Village Memorial Park Cemetery in Los Angeles. His widow died on November 17, 2011 at the age of ninety-three; her passing was publicized with a large paid obituary in the The Los Angeles Times, page A46, on November 27, 2011.