A chemically defined medium (also known as synthetic medium) is a growth medium suitable for the in vitro cell culture of human or animal cells in which all of the chemical components are known. Standard cell culture media commonly consist of a basal medium supplemented with animal serum (such as fetal bovine serum, FBS) as a source of nutrients and other ill-defined factors. The technical disadvantages to using serum include its undefined nature, batch-to-batch variability in composition, and the risk of contamination.

There is a clear distinction between serum-based media and chemically defined media. Serum-based media may contain undefined animal-derived products such as serum (purified from blood), hydrolysates, growth factors, hormones, carrier proteins, and attachment factors. These undefined animal-derived products will contain complex contaminants, such as the lipid content of albumin. In contrast, chemically defined media require that all of the components must be identified and have their exact concentrations known. Therefore, a chemically defined medium must be entirely free of animal-derived components and cannot contain either fetal bovine serum, bovine serum or human serum. To achieve this, chemically defined media is commonly supplemented with recombinant versions of albumin and growth factors, usually derived from rice or E. coli, or synthetic chemical such as the polymer polyvinyl alcohol which can reproduce some of the functions of BSA/HSA.

The constituents of a chemically defined media include: a basal media (such as DMEM, F12, or RPMI 1640, containing amino acids, vitamins, inorganic salts, buffers, antioxidants and energy sources), which is supplemented with recombinant albumin, chemically defined lipids, recombinant insulin and/or zinc, recombinant transferrin or iron, selenium and an antioxidant thiol such as 2-mercaptoethanol or 1-thioglycerol. Chemically defined media that are designed for the cultivation of cells in suspension additionally contain suitable surfactants such as poloxamers in order to reduce shear stress caused by shaking and stirring.

Chemically defined media also allows researchers who are studying in the field of cell physiology (especially extracellular) and or molecule–cell interactions to eliminate any variables that may arise due to the effects of unknown components in the medium.

Animal culture media can be divided into six subsets based on the level of defined media (Jayme and Smith, 2000): From lowest definition to highest these are:

The term chemically defined media is often misused in the literature to refer to serum albumin-containing media. The term 'defined media' can also be used to describe this type of media. Media formulations containing the media supplement B27 (supplied by Invitrogen) are often erroneously referred to as chemically defined media (e.g. Yao et al., 2006) despite this product containing bovine serum albumin (Chen et al., 2008) using the above definitions this type of media is referred to as serum-free media. Peptide-free, protein-free, chemically defined media are rarely successfully formulated except for CHO and insect cells.

Animal protein-free media, containing human serum albumin, human transferrin, but potentially animal-derived insulin and lipids.

Xeno-free media, containing human serum albumin, human transferrin, human insulin, and chemically defined lipids.