Magdalene's origins reach back to 1428, when a Benedictine hostel was established on the site. It passed through several incarnations — including a period under the patronage of the Duke of Buckingham, when it was known as Buckingham College — before Thomas Audley, Lord Chancellor under Henry VIII, refounded it in 1542 as the College of St Mary Magdalene. The transaction that secured the college's land was, according to later legal analysis, "almost certainly illegal." An agent named Spinola persuaded the master and fellows to accept an annual rental increase in exchange for a valuable London property. The college immediately pursued the matter in court. In 1615, master Barnaby Goche took up the legal fight so aggressively that he and the senior fellow were imprisoned for two years. When the Quayside development adjacent to the college was completed in 1989, a gargoyle of Spinola was installed, spitting water into the Cam. The college had waited 350 years for that particular revenge.

The most famous alumnus of Magdalene is Samuel Pepys, the 17th-century naval administrator and diarist whose journals record London life from 1660 to 1669 in extraordinary detail — the Great Fire, the plague, the court of Charles II, his own anxieties and pleasures — in a shorthand cipher that remained largely unread until Mynors Bright deciphered it in the 19th century. Pepys donated his library of 3,000 volumes to Magdalene upon his death, along with the six bookcases he had made to house them, in exactly the order he arranged them. The Pepys Library still exists in Second Court, the books still in their cases, the cases in their original arrangement. It is one of the best-preserved 17th-century private libraries in England. A portrait of Pepys by Peter Lely hangs in the college hall, watching over the candlelit dinners below.

C.S. Lewis was a fellow of Magdalene from 1954 until his death in 1963, having spent the previous thirty years at Oxford. He came to Cambridge as the first holder of the Chair of Medieval and Renaissance Literature. Lewis's time at Magdalene was quieter than his Oxford years, but no less productive — he wrote parts of his later fiction and theological works here. The chair he held was later occupied by three other Magdalene fellows: J.A.W. Bennett, John Stevens, and Helen Cooper. George Mallory, whose famous answer to why he wanted to climb Everest — "Because it's there" — echoes across a century, was also a Magdalene man. One of the college's residential courts is named after him. John Gurdon, former master and Nobel Prize winner in 2012 for his work on cellular reprogramming, is among the college's honorary fellows.

Cambridge colleges began admitting women in 1972, when Churchill, Clare, and King's led the way. By 1985, Oriel College, Oxford, had followed — leaving Magdalene as the last all-male Oxbridge college. The decision to change came in 1986. Women joined the college in 1988. Some male undergraduates responded by wearing black armbands and flying the college flag at half-mast. The photograph of that protest circulated widely. Within a generation, Magdalene's student body had become evenly mixed in terms of sex, race, and educational background. The college's close affiliation with international scholarships has also brought a strong presence from Southeast Asia. Its new library, designed by Niall McLaughlin Architects and opened in 2022, won the RIBA Stirling Prize — the first Oxbridge building to receive that accolade. The building that symbolizes Magdalene's future looks out over the Fellows' Garden and the River Cam.

The dining hall at Magdalene was built in the early 16th century. Unlike every other hall in Oxbridge, it has never been connected to gas or electric lighting. Formal dinners are served by candlelight — not as an affectation but because the hall was never renovated to accommodate anything else. The walls are hung with 15 portraits of benefactors and former members. A double staircase leads to a minstrels' gallery. The High Table stands on a platform one step above the students' long benches. Formal hall is held every evening; it is one of the traditions Magdalene is most known for, along with its candlelit aesthetic. Across Second Court, Bright's Building commemorates Mynors Bright, the man who deciphered Pepys. Above the entrance to one staircase of the Lutyens building — designed by Sir Edwin Lutyens in the 1930s — the crest of Harvard appears, a reminder that Henry Dunster studied at Magdalene in the 1620s before emigrating to Massachusetts and helping found Harvard University.