Born in Washington DC, Ramsey was educated at Columbia and at Harvard, where he obtained his PhD. He has served as professor of physics at Harvard from 1947 until his retirement in 1986.

Ramsey was a student of Isidor Rabi and worked with him on the rotational magnetic moments of molecules, showing how these depend on the mass of the nuclei. During World War II he worked first on radar, and later at the Los Alamos Laboratory. His subsequent work was on both high-energy particle scattering and on low-energy magnetic resonance.

In 1947 he began work on a new, more accurate, molecular-beam resonance technique using two separate radiofrequency fields. This was used to measure nuclear magnetic moments and nuclear quadrupole moments and also to investigate the magnetic interactions within simple molecules. Ramsey used the idea of magnetic shielding to interpret the chemical shifts found in nuclear magnetic resonance spectra.

Ramsey, along with D. Kleppner and H. M. Goldenberg, also developed the hydrogen maser. This used a molecular beam of hydrogen atoms and depended for its action on the hyperfine splitting of energy levels (splitting caused by interaction of electron energy levels with the nuclear magnetic moment). It was a highly accurate device, capable of measuring frequency to an accuracy of 1 part in 10¹².

Ramsey also worked on the thermodynamics and statistical mechanics of systems at low temperatures, pointing out that there is a possibility of certain nuclear spin systems having a negative thermodynamic temperature (i.e., a temperature below absolute zero).

For his work on molecular beams Ramsey was awarded the 1989 Nobel Prize for physics, which he shared with H. Dehmelt and W. Paul. He is also the author of a standard work on the subject, Molecular Beams (Oxford, 1963).