The Lord Chamberlain's Men was an English company of actors, or a "playing company" (as it then would likely have been described), for which William Shakespeare wrote during most of his career. Richard Burbage played most of the lead roles, including Hamlet, Othello, King Lear, and Macbeth. Formed at the end of a period of flux in the theatrical world of London, it had become, by 1603, one of the two leading companies of the city and was subsequently patronized by James I.

It was founded during the reign of Elizabeth I of England in 1594 under the patronage of Henry Carey, 1st Baron Hunsdon, then the Lord Chamberlain, who was in charge of court entertainments. After Carey's death on 23 July 1596, the company came under the patronage of his son, George Carey, 2nd Baron Hunsdon, for whom it was briefly known as Lord Hunsdon's Men. When George Carey in turn became Lord Chamberlain on 17 March 1597, it reverted to its previous name. The company became the King's Men in 1603 when King James ascended the throne and became the company's patron. The company held exclusive rights to perform Shakespeare's plays.

From 1594 the players performed at The Theatre, in Shoreditch. Problems with the landlord caused the company to move to the nearby Curtain Theatre in 1597. On the night of 29 December 1598, The Theatre was dismantled by the Burbage brothers, along with William Smith, their financial backer, Peter Street, a carpenter, and ten to twelve workmen. The beams were then carried south of the river to Southwark to form part of their new playhouse, the Globe Theatre. Built in 1599, this theatre was destroyed in a fire on 29 June 1613. The Globe was rebuilt by June 1614 and finally closed in 1642. The company also toured Britain, and visited France and Belgium.

A modern reconstruction of the original Globe, named "Shakespeare's Globe", was opened in 1997 near the site of the original theatre.

The initial form of the Chamberlain's men arose largely from the departure of Edward Alleyn from Lord Strange's Men and the subsequent death of Lord Strange himself, in the spring of 1594. Yet the ultimate success of the company was largely determined by the Burbage family. James Burbage was the impresario who assembled the company and directed its activities until his death in 1597; his sons Richard and Cuthbert were members of the company, though Cuthbert did not act. This connection with the Burbages makes the Chamberlain's Men the central link in a chain that extends from the beginning of professional theatre (in 1574, James Burbage led the first group of actors to be protected under the 1572 statute against rogues and vagabonds) in Renaissance London to its end (in 1642, the King's Men were among the acting companies whose activities were ended by Parliament's prohibition of the stage.)

The Chamberlain's Men comprised a core of eight "sharers", who split profits and debts; perhaps an equal number of hired men who acted minor and doubled parts; and a slightly smaller number of boy players, who were sometimes bound apprentices to an adult actor. The original sharers in the Chamberlain's were eight. Probably the most famous in the 1590s to the 1600s was William Kempe, who had been in the company of the Earl of Leicester in the 1580s, and had later joined the King's Men. As the company's clown, he presumably took the broadest comic role in every play; he is identified with Peter in the quarto of Romeo and Juliet, and probably also originated Dog-berry in Much Ado About Nothing and Bottom in A Midsummer Night's Dream. Kempe has traditionally been viewed as the object of Hamlet's complaint about extemporising clowns; whether this association is right or wrong, Kempe had left the company by 1601. Another two sharers from Strange's Men had a long-standing association with Kempe. George Bryan had been in Leicester's Men in the 1580s, and at Elsinore with Kempe in 1586; because he is not mentioned in later Chamberlain's or King's Men documents, it is assumed that Bryan retired from the stage in 1597 or 1598. (Bryan lived on for some years; in the reign of James, he is listed as a Groom of the Chamber, with household duties, as late as 1613.) Thomas Pope, another Leicester's veteran, retired in 1600 and died in 1603. Both Bryan and Pope came to the company from Lord Strange's Men. Augustine Phillips also came from Strange's Men. He remained with the troupe until his death in 1605.

Two younger actors who came from Strange's, Henry Condell and John Heminges, are most famous now for collecting and editing the plays of Shakespeare's First Folio (1623). Both were relatively young in 1594, and both remained with the company until after the death of King James; their presence provided an element of continuity across decades of changing taste and commercial uncertainty.

(Some scholars have theorised that the company maintained its original eight-sharer structure, and that as any man left, through retirement or death, his place as sharer was filled by someone else. So, Bryan was replaced by William Sly, ca. 1597; Kempe was replaced by Robert Armin, ca. 1599; Pope was replaced by Condell, ca. 1600. But this scheme, while possible, is not proven by the available evidence.)