A biosignature (sometimes called chemical fossil or molecular fossil) is any substance – such as an element, isotope, molecule, or phenomenon – that provides scientific evidence of past or present life on a planet or moon. Measurable attributes of life include its physical or chemical structures, its use of free energy, and the production of biomass and wastes.

The field of astrobiology uses biosignatures as evidence for the search for past or present extraterrestrial life. Candidate biosignatures strongly indicate some of the earliest known life forms, aid studies of the origin of life on Earth as well as the possibility of life on Mars, Venus and elsewhere in the universe.

Biosignatures can be grouped into ten broad categories:

Determining whether an observed feature is a true biosignature is complex. There are three criteria that a potential biosignature must meet to be considered viable for further research: Reliability, survivability, and detectability.

A biosignature must be able to dominate over all other processes that may produce similar physical, spectral, and chemical features. When investigating a potential biosignature, scientists must carefully consider all other possible origins of the biosignature in question. Many forms of life are known to mimic geochemical reactions. One of the theories on the origin of life involves molecules developing the ability to catalyse geochemical reactions to exploit the energy being released by them. These are some of the earliest known metabolisms (see methanogenesis). In such case, scientists might search for a disequilibrium in the geochemical cycle, which would point to a reaction happening more or less often than it should. A disequilibrium such as this could be interpreted as an indication of life.

A biosignature must be able to last for long enough so that a probe, telescope, or human can be able to detect it. A consequence of a biological organism's use of metabolic reactions for energy is the production of metabolic waste. In addition, the structure of an organism can be preserved as a fossil and we know that some fossils on Earth are as old as 3.5 billion years. These byproducts can make excellent biosignatures since they provide direct evidence for life. However, in order to be a viable biosignature, a byproduct must subsequently remain intact so that scientists may discover it.

A biosignature must be detectable with the most latest technology to be relevant in scientific investigation. This seems to be an obvious statement, however, there are many scenarios in which life may be present on a planet yet remain undetectable because of human-caused limitations.

Every possible biosignature is associated with its own set of unique false positive mechanisms or non-biological processes that can mimic the detectable feature of a biosignature. An important example is using oxygen as a biosignature. On Earth, the majority of life is centred around oxygen. It is a byproduct of photosynthesis and is subsequently used by other life forms to breathe. Oxygen is also readily detectable in spectra, with multiple bands across a relatively wide wavelength range, therefore, it makes a very good biosignature. However, finding oxygen alone in a planet's atmosphere is not enough to confirm a biosignature because of the false-positive mechanisms associated with it. One possibility is that oxygen can build up abiotically via photolysis if there is a low inventory of non-condensable gasses or if the planet loses a lot of water. Finding and distinguishing a biosignature from its potential false-positive mechanisms is one of the most complicated parts of testing for viability because it relies on human ingenuity to break an abiotic-biological degeneracy, if nature allows.