In 1934–36, Berle was heard regularly on The Rudy Vallee Hour, and he got much publicity as a regular on The Gillette Original Community Sing, a Sunday night comedy-variety program broadcast on CBS from September 6, 1936 to August 29, 1937. In 1939, he was the host of Stop Me If You've Heard This One with panelists spontaneously finishing jokes sent in by listeners.

Three Ring Time, a comedy-variety show sponsored by Ballantine Ale, was followed by a 1943 program sponsored by Campbell's Soups. The audience participation show Let Yourself Go (1944–1945) could best be described as slapstick radio with studio audience members acting out long suppressed urges (often directed at host Berle). Kiss and Make Up, on CBS in 1946, featured the problems of contestants decided by a jury from the studio audience with Berle as the judge. He also made guest appearances on many comedy-variety radio programs during the 1930s and 1940s.

Scripted by Hal Block and Martin Ragaway, The Milton Berle Show brought Berle together with Arnold Stang, later a familiar face as Berle's TV sidekick. Others in the cast were Pert Kelton, Mary Schipp, Jack Albertson, Arthur Q. Bryan, Ed Begley and announcer Frank Gallop. Sponsored by Philip Morris, it aired on NBC from March 11, 1947 until April 13, 1948.

His last radio series was The Texaco Star Theater, which began September 22, 1948 on ABC and continued until June 15, 1949, with Berle heading the cast of Stang, Kelton and Gallop, along with Charles Irving, Kay Armen, and double-talk specialist Al Kelly. It employed top comedy writers (Nat Hiken, brothers Danny and Neil Simon, Leo Fuld, Aaron Ruben), and Berle later recalled this series as "the best radio show I ever did... a hell of a funny variety show". It served as a springboard for Berle's rise as television's first major star.