The dyslexia continued to hamper her in her search for a career. Young Susan Hampshire originally wanted to be a nurse but found it impossible, and she found the theatre in her teens.

Susan Hampshire's first film appearance was in the movie The Woman in the Hall. She decided to become an actress as a child and worked in a theatre before moving on to film and television work.

Hampshire first became famous after playing the lead in a 1962 BBC adaptation of What Katy Did. Soon afterwards, she was taken up by Walt Disney, and starred in The Three Lives of Thomasina (opposite Patrick McGoohan) and The Fighting Prince of Donegal. She would later appear opposite McGoohan in two episodes of Danger Man. Later, she portrayed conservationist Joy Adamson in the 1972 film Living Free, sequel to the 1966 classic Born Free. She has also ventured into musicals, starring opposite Cliff Richard, in Wonderful Life, in Follow That Girl and more recently (1991) in The King and I.

Susan Hampshire is best known for her work on television. She appeared in several popular television serials, notably in the BBC's blockbuster, The Forsyte Saga (1967), in which she played Fleur. The popularity of this series was a factor in the creation of the PBS program Masterpiece Theatre (in which the first series aired was The First Churchills in which she played Sarah Churchill). 1973 saw Hampshire on US television with Kirk Douglas in a musical version of Dr. Jekyll and Mr. Hyde. Her most recent TV role was as Molly Macdonald, in the highly popular Monarch of the Glen (2000–2006).

Miss Hampshire received Emmy Awards from the Academy of Television Arts & Sciences for her roles in The Forsyte Saga (1970), The First Churchills (1971), and Vanity Fair (1973). Other miniseries in which she appeared are The Pallisers, The Barchester Chronicles, and Coming Home.

Susan Hampshire has been active on the stage over the years, taking the lead roles in many leading plays, such as Relative Values, Lady in the Van, The Importance of Being Earnest and For no good Reason, a play written by Nathalie Sarraute in which she appears with Susannah York under the direction of French director Simone Benmussa. In 2007 she was in a ground breaking play The Bargain, based on a meeting between Robert Maxwell and Mother Teresa. She even ventured into the British pantomime tradition, playing the Fairy Godmother at the New Wimbledon Theatre in 2005–2006 and at the New Victoria Theatre in Woking in 2006–2007.

In 2008 she joined the relatively small band of actors who have played two generations in the same play on different occasions. Her appearance at Chichester Festival Theatre in Somerset Maugham's "The Circle" as Lady Catherine Champion-Cheney in Summer 2008 followed on from her appearance in the same play (and venue) as Elizabeth Champion-Cheney (Lady Catherine's daughter-in-law) somewhat earlier in her career.

Until the publication of her autobiography, Susan's Story, few people were aware of her struggle with dyslexia. She was undiagnosed until she was an adult and since has become a prominent campaigner in the UK on dyslexia issues. Her book on dyslexia, "Every Letter Counts", was highly acclaimed. In 1995, she was appointed an OBE (Officer of the Order of the British Empire) in connection with that work.

Susan Hampshire has also published a book called The Maternal Instinct, about women and fertility issues. She herself suffered a number of miscarriages over the years.

She has written children's books, including "Lucy Jane at the Ballet", "Lucy Jane and the Russian Ballet", "Lucy Jane and the Dancing Competition", "Lucy Jane on Television", "Bear's Christmas", "Rosie's First Ballet Lesson", and "Rosie's Ballet Slippers", and various books and videos about her lifelong hobby gardening, including "Easy Gardening", "My Secret Garden", and "Trouble Free Gardening".