Loretta Mary Aiken was born in Brevard, North Carolina on March 19, 1894 to James Aiken and Mary Smith, who married on May 21, 1891, in Transylvania County, North Carolina. Loretta was one of 16 children.

Her father owned and operated several successful businesses, while her mother kept house and took in boarders. While working as a volunteer fireman in 1909, her father died when a fire engine exploded. Loretta was 15 years old at the time. In 1910, her mother took over their primary Business, a general store. She was killed after being run over by a truck while returning home from church on Christmas Day.

She came out as a lesbian at the age of twenty-seven, becoming one of the first openly gay comedians. During the 1920s and 1930s she appeared in androgynous clothing (as she did in the film version of The Emperor Jones with Paul Robeson) and recorded several of her early "lesbian stand-up" routines.

In the 1960s, she became known to a wider white audience, playing Carnegie Hall in 1962, and making a number of mainstream TV appearances, particularly her multiple appearances on The Smothers Brothers Comedy Hour when that CBS show was number one on television in the late 1960s, which introduced her to a whole new Boomer audience.

She also added the occasional satirical song to her jokes, and her (completely serious and melancholy) cover version of "Abraham, Martin and John" hit #35 on the Billboard Hot 100 on July 19, 1969. At 75 years old, Mabley became the oldest living person ever to have a US Top 40 hit (Louis Armstrong, who would have been 86 when "What a Wonderful World" became a hit in 1988, is the oldest overall, although Armstrong was younger than Mabley when the record was made).

Loretta Aiken took her stage name, Jackie Mabley, from an early boyfriend, commenting to Ebony in a 1970s interview that he had taken so much from her, it was the least she could do to take his name. Later she became known as "Moms" because she was indeed a "Mom" to many other comedians on the circuit in the 1950s and 1960s.

Mabley had six children: Bonnie, Christine, Charles, and Yvonne Ailey, and two given up for adoption when she was a teenager. She died from heart failure in White Plains, New York on May 23, 1975. She is interred at Ferncliff Cemetery, Hartsdale, New York.

Mabley was the inspiration for the character of Grandma Klump in The Nutty Professor. She is the subject of Whoopi Goldberg Presents Moms Mabley, a documentary film which first aired on HBO on November 18, 2013.

This documentary was nominated for two Creative Arts Emmy Awards at the 66th ceremony held on August 16, 2014, at the Nokia Theatre in Downtown Los Angeles: Outstanding Documentary or Nonfiction Special and Outstanding Narrator for Whoopi Goldberg. In 2015, she was named by Equality Forum as one of their 31 Icons of the 2015 LGBT History Month.

Mabley was featured during the "HerStory" video tribute to notable women on U2's tour in 2017 for the 30th anniversary of The Joshua Tree during a performance of "Ultraviolet (Light My Way)" from the band's 1991 album Achtung Baby.