Nicholas Hilliard, the son of Richard Hilliard (1519–1594) of Exeter, Devon, England, a staunchly Protestant goldsmith who was high sheriff of the city and county in 1560, and Laurence, daughter of John Wall, a London goldsmith, may have been a close relative of Grace Hiller (Hilliar), first wife of Theophilus Eaton (1590–1657), the co-founder of New Haven Colony in America.

He appears to have been attached at a young age to the household of the leading Exeter Protestant John Bodley, the father of Thomas Bodley who founded the Bodleian Library in Oxford.

John Bodley went into exile on the accession of the Catholic Queen Mary I of England, and on May 8, 1557 Hilliard, then ten years old, was recorded in Geneva as one of an eleven-strong Bodley family group at a Calvinist service presided over by John Knox.

Calvinism does not seem to have struck with Hilliard, but the fluent French he had acquired abroad will prove useful.

Thomas Bodley, two years older, continued an intensive classical education under leading scholars in Geneva, but it is not clear to what extent Hilliard was given similar studies.

Hilliard had painted a portrait of himself at the age of thirteen in 1560 and is said to have executed one of Mary Queen of Scots when he was eighteen years old.

Apprenticed to the goldsmith Robert Brandon in 1562, Hilliard had emerged from his apprenticeship at a time when a new royal portrait painter was "desperately needed".

Two panel portraits long attributed to him, the "Phoenix" and "Pelican" portraits, are dated to about 1572-76.

The first true English miniature painter born in England, Hilliard had been appointed limner (miniaturist) and goldsmith to Elizabeth I at an unknown date; his first known miniature of the Queen is dated 1572, and already in 1573 he is granted the reversion of a lease by the Queen for his "good, true and loyal service."

He had made "a booke of portraitures" in 1571 for the Earl of Leicester, the Queen's favorite, which is likely to be how he became known to the Court; several of his children are to be named after Leicester and his circle.

The goldsmith and miniaturist Nicholas Hilliard, recently married, had left in 1576 for France, despite the royal patronage, "with no other intent than to increase his knowledge by this voyage, and upon hope to get a piece of money of the lords and ladies here for his better maintenance in England at his return", carefully reported the English Ambassador in Paris, Sir Amyas Paulet, with whom Hilliard is to stay for much of the time.

Francis Bacon is attached to the embassy, and Hilliard had done a miniature of him in Paris.

He appears in the papers of the duc d'Alençon, a suitor of Queen Elizabeth, under the name of "Nicholas Belliart, peintre anglois" in 1577, receiving a stipend of two hundred livres.

Hilliard remains at the French court until 1578-79, mixing in the artistic circles round the court, staying with Germain Pilon and George of Ghent, respectively the Queen's sculptor and painter, and meeting Ronsard, who perhaps paid him the rather double-edged compliment later quoted by Hilliard: "the islands indeed seldom bring forth any cunning man, but when they do it is in high perfection".

The miniature of Madame de Sourdis, certainly the work of Hilliard, is dated 1577, in which year she was a maid of honor at the French court; and other portraits which are his work are believed to represent Gabrielle d'Estrées (niece of Madame de Sourdis), la princesse de Condé, and Madame de Montgomery.

The Earl of Mar, after a brief seizure of Stirling Castle in the hope of prompting English intervention, is forced to flee to England, where he receives the backing of Queen Elizabeth.

Hilliard designs and engraves Elizabeth’s second Great Seal of England in 1584.

The oldest extant accounts of Madog Ab Owain Gwynedd, the legendary twelfth century voyager to America, are in Richard Hakluyt's Voyages (1582) and David Powel's The Historie of Cambria (1584).

Hakluyt believed Madog had landed in Florida.