Educated in England, Eliot graduated from Jesus College, Cambridge, in 1622 and emigrated to Boston in 1631. From 1632 to his death he was pastor of the church at nearby Roxbury. With the support of his congregation and fellow ministers, he began a mission to the Native Americans, preaching at Nonantun (Newton) and at other towns. Groups of “praying Indians” soon arose, and by 1674 there were 14 villages with 4,000 converts. The following year, however, the communities suffered serious setbacks from persecutions that occurred during King Philip's War, and the villages never fully recovered.

Eliot's work was financed chiefly from England, where his activities inspired the creation of the Company for Propagating the Gospel in New England and Parts Adjacent in North America (1649). This was the first genuine missionary (mission) society. Eliot's methods set the pattern of subsequent “Indian missions” for almost two centuries. Civilization, he believed, was closely bound up with evangelization. His converts were gathered into Christian towns, governed by a biblical code of laws, and gradually introduced to the English manner of life. Each village had a school where the Indians were taught English and the handicrafts by which they could support themselves. After severe testing, believers were organized by covenant into a Puritan “church-state,” and native teachers and evangelists were trained. Eliot himself, called the “Apostle to the Indians,” produced the needed literature in the Massachusets (Massachuset) Algonquian language, beginning with his primer or catechism of 1654. His translation of the New Testament appeared in 1661, the Old Testament in 1663. Among his other works are The Christian Commonwealth (1659) and The Harmony of the Gospels (1678).