Isotope separation is the process of concentrating specific isotopes of a chemical element by removing other isotopes. The use of the nuclides produced is varied. The largest variety is used in research. By tonnage, separating natural uranium into enriched uranium and depleted uranium is the largest application. In the following text, mainly uranium enrichment is considered. This process is crucial in the manufacture of uranium fuel for nuclear power plants, and is also required for the creation of uranium-based nuclear weapons. Plutonium-based weapons use plutonium produced in a nuclear reactor, which must be operated in such a way as to produce plutonium already of suitable isotopic mix or grade.

Isotopes are distinct nuclear species of the same chemical element. They have the same atomic number and position in the periodic table, but differ in nucleon numbers due to different numbers of neutrons in their nuclei. While all isotopes of a given element have similar chemical properties, they have different atomic masses and physical properties.