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Cory Buxton and Eugene Provenzo place Mr. Wizard in a 19th-century tradition of "hands-on kitchen science" associated with Michael Faraday's popular science lectures and Arthur Good's collection of experiments for children, La Science Amusante (1893). In turn, LaFollette has written on the legacy of Herbert and other early innovators of science television, "Production approaches that are now standard practice on NOVA and the Discovery Channel derive, in fact, from experimentation by television pioneers like Lynn Poole and Don Herbert and such programs as Adventure, Zoo Parade, Science in Action, and the Bell Telephone System’s science specials. These early efforts were also influenced by television’s love of the dramatic, refined during its first decade and continuing to shape news and public affairs programming, as well as fiction and fantasy, today."

In the mid-1950s, Herbert also appeared on the General Electric Theater as the "General Electric Progress Reporter" and would introduce spokesman Ronald Reagan and his family to the viewing audience. In some episodes, he would appear alongside Reagan and demonstrate to the audience how General Electric was helping people to, "Live better electrically."

After the war Herbert worked at a radio station in Chicago where he acted in children's programs such as the documentary health series It's Your Life (1949). It was during this time that Herbert formulated the idea of Mr. Wizard and a general science experiments show that used the new medium of television. Herbert's idea was accepted by Chicago NBC station WNBQ and the series Watch Mr. Wizard premiered on March 3, 1951. The weekly half-hour live television show, co-produced by Jules Power, featured Herbert as Mr. Wizard and either a boy or a girl with whom Herbert performed interesting science experiments. The experiments, many of which seemed impossible at first glance, were usually simple enough to be re-created by viewers.

After Watch Mr. Wizard was cancelled in 1965, Herbert produced eight films in a series titled Experiment: The Story of a Scientific Search; these aired on public television in 1966. In the same year, Herbert produced the Science 20 series, which were 20-minute films of experiments that were designed for classroom use; a student would record and analyze data based on the film. In 1977, he began producing a series of How About episodes about scientific topics. These were 90-second films that could be used in news programs; by 1986, he produced 536 films.

In 1969, Herbert opened a Mr. Wizard Science Center in Wellesley, Massachusetts; the center no longer exists.

In 1982, Don Herbert was a guest on the first episode of Late Night with David Letterman.

In 1983, Herbert developed Mr. Wizard's World, a faster-paced version of his show that aired three times per week on the cable channel Nickelodeon. The show ran until 1990 and reruns were shown until 2000.

In 1993 children's science show Beakman's World paid homage to Herbert by naming its two penguin puppet characters "Don" and "Herb" after him.

In 1994, Herbert developed another new series of 15-minute spots called Teacher to Teacher with Mr. Wizard. The spots highlighted individual elementary science teachers and their projects. The series was sponsored by the National Science Foundation and was shown on Nickelodeon.

Herbert died June 12, 2007, of multiple myeloma, four weeks before what would have been his 90th birthday, at his home in Bell Canyon, California. In Herbert's obituary, Bill Nye wrote, "If any of you reading now have been surprised and happy to learn a few things about science watching "Bill Nye the Science Guy," keep in mind, it all started with Don Herbert." Jamie Hyneman and Adam Savage, principals of the television program MythBusters (2002–2016), have been described as being "reverent" of Herbert's work as Mr. Wizard. Five months after Herbert died, MythBusters aired a two-hour episode entitled "Special Super-sized Myths" "Dedicated to Mr. Wizard".

Born in Waconia, Minnesota, Herbert was a general science and English major at the University of Wisconsin–La Crosse (then called La Crosse State Teachers College) who was interested in drama. His career as an actor was interrupted by World War II when he enlisted in the United States Army as a Private. Herbert later joined the United States Army Air Forces, took pilot training, and became a B-24 bomber pilot who flew 56 combat missions from Italy with the 767th Bomb Squadron, 461st Bomb Group of the Fifteenth Air Force. When Herbert was discharged in 1945 he was a Captain and had earned the Distinguished Flying Cross and the Air Medal with three oak leaf clusters.