As a young man, Standish fought in the Netherlands, where he probably met the English religious exiles who later became known as the Pilgrims. He sailed with them to America on the “Mayflower” in 1620, serving as their military leader when they established Plymouth in New England. He learned the language of the local Indians and led several expeditions against hostile tribes. In 1627 he was a leader of the group in the colony that bought out the London investors. In 1628 Standish helped to break up the colony of Thomas Morton at nearby Merry Mount when it proved too unpuritanical to suit Plymouth. He also served as assistant governor and as treasurer of the Plymouth colony (1644–49). He moved to Duxbury in 1631 and remained there until his death.

There is no historical evidence for the story that he asked John Alden to propose marriage for him to Priscilla Mullins, as told in Longfellow's poem The Courtship of Miles Standish.