Born Frederick Llewellyn March in Dublin, Bartholomew was abandoned by his parents while a baby, and was raised in London by his aunt, whose name he took. While visiting the United States, Bartholomew was reportedly seen by film producer David O. Selznick who was soon to film Charles Dickens' David Copperfield (1935). Selznick had already cast an American boy in the role, but after meeting Bartholomew realised that the character would benefit from being played by a British actor. The all-star film was a success and Bartholomew was cast in a succession of prestigious film productions with some of the most popular stars of the day.

Among his successes of the 1930s were Anna Karenina (1935), with Greta Garbo and Fredric March, Professional Soldier (1935) with Gloria Stuart, Little Lord Fauntleroy (1936) with Dolores Costello, Lloyds of London (1937) with Madeleine Carroll and Tyrone Power, and Captains Courageous (1937) with Spencer Tracy.

By this time Bartholomew's success and level of fame had caused his parents to attempt to gain custody of him. A protracted legal battle saw much of the wealth Bartholomew had amassed spent on legal fees. He continued acting into the 1940s but was much less popular as a teenaged actor, and by the early 1950s had retired from film.

He established a career in advertising and distanced himself from Hollywood. Bartholomew was said to have been bitter over his lost fortune and his experiences in Hollywood, but by the early 1980s he was working as a producer for the soap opera As The World Turns. Shortly before his death he allowed an interview for the television documentary MGM: When the Lion Roars (1992).

He died from emphysema in Sarasota, Florida.

Freddie Bartholomew has a star on the Hollywood Walk of Fame for his contribution to motion pictures, at 6667 Hollywood Boulevard.