The foundations of mass spectrometry lie in the work of Thomson and Aston at the Cavendish Laboratories, Cambridge University. From 1897, the work carried out by Thomson and his co-workers receive 7 Nobel prizes in Physics and Chemistry. Thomson's original work on the existence and properties of canal rays (positive ions) was taken up by Aston and by the end of the First World War he had demonstrated the existence of several isotopes of non-radioactive elements. Aston used electrostatic and magnetic fields to separated isotope ions by their masses and focus them onto a photographic plate. Over the next few years a number of key names took up the early development of mass spectrometry, including Dempster, Herzog, Bainbridge and Nier. By the end of the 1930's mass spectrometry had become an established technique for the separation of atomic ions by mass.‌

In the 1940's the applications of mass spectrometry began to spread away from the previous mostly academic work into more practical fields like nuclear isotope enrichment and the analysis of the components of petroleum. The World's first commercial instrument became available in 1948 (The MS-2 - marketed by Vickers in Manchester, England). The MS-2 made use of EI ionisation and (by today's standards) had an extremely limited mass range of about 300 Da at limited resolution.

In the early 1950's, the fragmentation of small organic molecules was beginning to be understood, but the mass spectrometer was still extremely limited by mass and resolution. At this time, time-of-flight (TOF) analysis (Wiley and Maclaren) and quadruple analysis (Paul) were conceived. These early instruments were the forerunners of today's benchtop instruments seen in just about every chemical and biochemical lab in the world.

The next major breakthroughs were the coupling of gas and liquid chromatography to mass spectrometry. This allowed, for the first time, the analysis of mixtures of analytes without laborious separation by hand. The development of GC-MS was the trigger for the development of modern mass spectrometry. In 1956, the first biologically important molecules were successfully analysed. New ionisation techniques developed over the last 25 years (fast particle desorption, electrospray ionisation and matrix-assisted laser desorption/ionsation) have opened up the world of biological chemistry to mass spectrometry. Just about every compound class can be analysed by some sort of mass spectrometry (see the theory section for a description of the most important techniques) and the present mass records extend well into the megadalton (millions of mass) range.