The poem was Chapman's first significant literary work; it is furnished with abundant notes and references to classical Greek and Roman authors (41 in total, drawn from the Mythologiae of Niccolo Conti).
The three noblemen and their associates, like Sir Walter Raleigh, John Dee and Thomas Harriot among others, have been interpreted as members of a clique of advanced thinkers called The School of Night, who were interested in promoting new ideas like the Copernican and Galilean view of a heliocentric solar system, and the spirit of open inquiry that underlay it.
In the poem's second half, the portrait of the moon goddess Cynthia represents Queen Elizabeth I, and comprises the type of hagiographic personification that was common in the later Elizabethan era.
The poem appears to contain an autobiographical reference, indicating that Chapman served in a military campaign in the Dutch Republic under Sir Francis Vere, in the 1591-2 period.
There Vere helped defeat the Spanish force under the Duke of Parma:[5] Sent out in hast against some English force From statelie sited sconce-torn Nimigan Vnder whose walles the Wall most Cynthian Stretch her siluer limms loded with wealth Hearing our horse were marching downe by stealth[6] The modern composer Harrison Birtwistle composed an orchestral piece (2001) which he titled The Shadow of the Night after Chapman's poem.