Ameche was married to Honore Prendergast from 1932 until her death in 1986. They had six children. One, Ron Ameche, owned a restaurant, "Ameche's Pumpernickel" in Coralville, Iowa. He had two daughters, Connie and Bonnie. Ameche's younger brother, Jim Ameche, was also a well-known actor. His brother Bert was an architect who worked for the U.S. Navy in Port Hueneme, California, and then the U.S. Postal Service in Los Angeles, California.
Ameche had done well in college dramatics at Marquette University, and when a lead actor for a stock company production of Excess Baggage did not turn up, a friend persuaded him to stand in for the missing actor. He enjoyed the experience and got a juvenile lead in Jerry For Short in New York, followed by a tour in vaudeville with Texas Guinan until she dropped him from the act, dismissing him as "too stiff". He made his film debut in 1935, and by the late 1930s, had established himself as a major actor in Hollywood. He appeared in such films as Alexander's Ragtime Band (1938), and as the title character in The Story of Alexander Graham Bell (1939). It led to the use of the word, "ameche", as slang for telephone in common catchphrases, as noted by Mike Kilen in the Iowa City Gazette (December 8, 1993): "The film prompted a generation to call people to the telephone with the phrase: 'You're wanted on the Ameche.'" In the 1940 film Go West, Groucho Marx proclaims, "Telephone? This is 1870, Don Ameche hasn't invented the telephone yet". While in the 1941 film Ball of Fire, Barbara Stanwyck's character discusses the "ameche" slang usage, "Do you know what this means: I'll get you on the Ameche."
In 1940, he was voted the 21st-most-popular star in Hollywood. In 1944 he reportedly earned $247,677 for 1943, making him the second highest earner at 20th Century Fox after Spyros Skouras. Ameche played so many roles based on real people that on one of his radio broadcasts, Fred Allen (who shares a birthday with Ameche) joked, "Pretty soon, Don Ameche will be playing Don Ameche." Soon afterwards, in It's in the Bag! (1945), which starred Allen, Ameche indeed played himself in a bit part.
Don Ameche continued to work on television and movies until his death from cancer in 1993...