Heyrovský, the son of an academic lawyer, was educated at Charles University in his native Prague and at University College, London. He joined the staff of Charles University in 1919 where he served as professor of physics from 1919 until 1954. From 1950 he was also head of the Central Polarographic Institute, which, since 1964, has borne his name.

Heyrovský is best known for his discovery and development of polarography, which he described in 1922. This is one of the most versatile of all analytical techniques. It depends on the fact that in electrolysis the ions are discharged at an electrode and, if the electrode is small, the current may be limited by the rate of movement of ions to the electrode surface. In polarography the cathode is a small drop of mercury (constantly forming and dropping to keep the surface clean). The voltage is increased slowly and the current plotted against voltage. The current increases in steps, each corresponding to a particular type of positive ion in the solution. The height of the steps indicates the concentration of the ion. For his work, Heyrovský was awarded the Nobel Prize for chemistry in 1959.