Born in Lancashire, UK, Macaulay moved to Bloomfield, New Jersey at the age of eleven. He began drawing while in the United States. After graduating from high school in Cumberland, Rhode Island in 1964, he enrolled in the Rhode Island School of Design (RISD), from which he received a bachelor's degree in architecture. He spent his fifth year at RISD in the European Honors Program, studying in Rome, Herculaneum and Pompeii.

Macaulay's books have sold more than two million copies in the United States, been translated into a dozen languages, and been widely praised. Time magazine said of his work, "What [Macaulay] draws he draws better than any other pen-and-ink illustrator in the world". His numerous awards include the MacArthur Fellows Program award, the Caldecott Medal, won for his book Black and White, the Boston Globe-Horn Book Award, the Christopher Award, an American Institute of Architects Medal, the Washington Children's Book Guild Nonfiction Award, the Deutscher Jugendliteraturpreis, and a Dutch Silver Slate Pencil Award. He was a two-time nominee for the Hans Christian Andersen Award and is the recipient of the Bradford Washburn Award, presented by the Museum of Science in Boston to an outstanding contributor to science.

In June 2007, his work was the subject of the exhibition "David Macaulay, the Art of Drawing Architecture" which opened for a limited run through January 2008 (later extended to May 4) at the National Building Museum in Washington, D.C.

Macaulay is the author of several books on architecture and design. His first book, Cathedral (1973), was a history, extensively illustrated with pen-and-ink drawings, of the construction of a fictitious but typical Gothic cathedral. This was followed by a series of books of the same type: City (1974), on the construction of Verbonia, a fictitious but typical Roman city; Pyramid (1975), on the building of monuments to the Egyptian Pharaohs; Castle (1977), on the construction of Aberwyvern castle, a fictitious but typical Medieval Castle; Mill (1983), on the evolution of New England mills; and Mosque (2003). Other books in this series are Underground (1976), which describes the building foundations and support structures (such as water and sewer pipes) that underlie a typical city intersection, and Unbuilding (1980), which describes the hypothetical dismantling of the Empire State Building in preparation for re-erection in the Middle East.

Macaulay has illustrated a number of other books, including the popular The Way Things Work (1988, text by Neil Ardley) which was expanded and rereleased as The New Way Things Work (1998). These works remain his most commercially successful. He has also written a number of children's fiction books.

His books often display a whimsical humor. Illustrations in The Way Things Work depict cave people and woolly mammoths operating giant-sized versions of the devices he is explaining. Motel of the Mysteries, written in 1979 following the 1976—1979 exhibition of the Tutankhamun relics in the USA, concerns the discovery by future archaeologists of an American motel and the archaeologists' ingenious interpretation of the motel and its contents as a funerary and temple complex. Baaa is set after the human race has somehow gone extinct. Sheep discover artifacts of lost human civilization and attempt to rebuild it. However, the new sheep-inhabited world develops the same side effects of economic disparity, crime, and war.

To research his book The Way We Work, Macaulay spent years talking and studying with doctors and researchers, attending medical procedures, and laboriously sketching and drawing. He worked with medical professionals such as Lois Smith (a professor at Harvard University and researcher at Children's Hospital Boston) and medical writer Richard Walker to ensure the accuracy of both his words and his illustrations. Anne Gilroy, clinical anatomist in the departments of surgery and cell biology at the University of Massachusetts Medical School, consulted on the book, and says of Macaulay, "His remarkable curiosity and meticulous research led him into some of the most complicated facets of the human body yet he tells this story with simplicity, ingenuity and humor."