1. What a culture medium is, in three points

A culture medium is the liquid that cells sit in so that they can survive, multiply and do their work outside the body. Inside the body, blood and tissue fluid deliver nutrients, salts and hormones. The easiest way to think of a medium is as an artificial formulation that recreates that supply (our commentary).

• The backbone: the foundation is a “basal medium”: water with inorganic salts, sugar, amino acids, vitamins and a buffer dissolved in it. In 1955 Harry Eagle identified the minimum set of components needed by mouse L cells and human HeLa cells, and this is regarded as the starting point for many of today's basal mediaSourced
• What gets added: most cells will not grow on a basal medium alone. The gap used to be filled by adding serum (fetal bovine serum and the like), but because serum is undefined and varies widely from lot to lot, it has increasingly been replaced by serum-free media that add the necessary factors one by oneSourced
• Why there are so many types: the best formulation differs with the cell (antibody-producing CHO cells, iPS cells, T cells, insect cells and so on), with the goal (expanding cells, differentiating them, or making them produce something) and with the culture mode (batch, fed-batch or perfusion) (our commentary)

2. What goes into it: components and their roles

First, split the contents of a medium into “the base” (the basal medium) and “what is added for the purpose”.

Sources for the “What the literature says” column: Yao and Asayama [Ref. 9], Eagle [Ref. 1], Good et al. [Ref. 32], Chen et al. [Ref. 14], Berthois et al. [Ref. 31]. The “Role” column is this article's general explanation. Conversions are our calculation (glucose 180.16 g/mol, phenol red 354.4 g/mol).

3. The family tree of basal media, 1950 to 1980

Many of the basal media in use today were born in the thirty years or so between 1950 and 1980. Each was created to grow a particular kind of cell.

All Sourced. Developers and years follow each original paper [Refs. 1 to 8, 10 and 11]; targets and features follow the review by Yao and Asayama [Ref. 9]; details of IMDM follow the original paper by Iscove and colleagues [Ref. 7]. The only original paper whose full text this article could read was the IMDM paper (Iscove et al.); the others were checked against their PubMed records and the review.

4. Our calculation: bicarbonate and CO2

Many media hold their pH through the combination of sodium hydrogen carbonate (bicarbonate) and the CO2 in the incubator. Using the Henderson–Hasselbalch equation, the review by Yao and Asayama shows that MEM (26 mmol/L bicarbonate) needs 5% CO2, Ham's F-12 (14 mmol/L) needs 2%, and DMEM (44 mmol/L) needs 10%Sourced. It adds that these are theoretical values, and in practice it is better to measure the pH after equilibration and fine-tune the CO2 levelSourced.

5. How serum is handled: five stages

“Serum-free”, “animal-component free”, “xeno-free”, “chemically defined”: media catalogues are full of these labels. Each one describes how far the medium has moved away from serum, a liquid whose content nobody fully knows.

What serum was doing

The review by Yao and Asayama summarises the functions of fetal bovine serum (FBS) as followsSourced.

• A source of amino acids, proteins, vitamins, sugars, lipids, hormones, growth factors, inorganic salts and trace elements
• Raising the pH buffering capacity of the medium
• Reducing shear stress (physical damage) from pipetting and stirring
• Changing the state of the culture surface so that adherent cells attach and grow more readily

Why people still want to stop using it

Sources: Yao and Asayama [Ref. 9], EMA guideline EMA/CHMP/BWP/457920/2012 rev 1 [Ref. 13], van der Valk et al. [Ref. 12]. The EMA guideline was adopted on 30 May 2013 and came into effect on 1 December 2013.

On inactivating viruses in serum, the EMA guideline notes that gamma irradiation is the most commonly used method, while also accepting other validated methodsSourced. Whether a process keeps serum or moves away from it, the underlying question is the same: how do you guarantee what is in a liquid raw material? (our commentary)

6. A materials engineer's view (1): albumin was working as an adsorbent

7. Media by purpose: antibodies, viral vectors, insect cells, T cells, iPS cells, differentiation, organoids

Starting from a basal medium, formulations branch according to which cell you are growing and what you want it to do. Below are representative examples that could be confirmed in primary sources.

(1) For producing antibodies and other proteins (CHO cells)

The review by Ritacco and colleagues describes how media for recombinant protein production in CHO cells moved from early formulations containing animal-derived and complex components to serum-free media and then chemically defined (CD) media, and then developed further towards optimising nutrient supply and limiting the build-up of waste products through fed-batch and perfusion cultureSourced. In 2015 a group from Chugai Pharmaceutical, noting that chemically defined media are currently used for fed-batch culture of antibody-producing CHO cells, reported an optimisation of feed mediaSourced.

(2) For producing viral vectors (HEK293 cells)

HEK293 cells, derived from human embryonic kidney, are widely used to manufacture the adeno-associated virus (AAV) vectors used in gene therapy. In 2016 Grieger and colleagues reported adapting adherent HEK293 cells from a clinical master cell bank to animal-component-free suspension culture and producing AAV vectors in shake flasks and a bag-type bioreactorSourced. One of the items they optimised was the choice of a serum-free suspension medium that supports both cell growth and transfection (gene delivery)Sourced. In other words, a medium for growing cells and a medium for getting genes into them are asked to do different things (our commentary).

(3) For insect cells

Insect cells are used, among other things, for protein production with baculovirus. In 1962 Grace reported in Nature that he had established four cell strains from insect tissueSourced. In 1977 Vaughn and colleagues reported establishing two cell lines from pupal tissue of the fall armyworm (Spodoptera frugiperda): one (IPLB-SF 21) in a medium supplemented with insect body fluid (haemolymph) and the other (IPLB-SF-1254) in a medium combining vertebrate serum with haemolymphSourced. Insect cell media come from a lineage separate from that of mammalian media (our commentary).

(4) For T cells, NK cells and other immune cells

Media for expanding T cells have included ones supplemented with human serum (the paper by Medvec and colleagues uses a human-serum medium as its comparator). In 2018 Medvec and colleagues reported developing a chemically defined medium that expands all T-cell subsets without human serumSourced. In a humanised mouse model, T cells expanded without serum controlled tumours for longerSourced. This is an experimental result in mice, however, and does not show any therapeutic effect in humans. For NK cell media, this article could not confirm original papers within the scope of its research, so they are not described separately.

(5) For maintaining pluripotent stem cells (ES and iPS cells)

Sources: Ludwig et al. [Refs. 15 and 16], STEMCELL Technologies product page [Ref. 17], Chen et al. [Ref. 14], Nakagawa et al. [Ref. 18], Kuo et al. [Ref. 35].

What Fig. 5 shows is that growth factors are indispensable even in minute amounts. TGFβ1 is present at 2 µg/L: two millionths of a gram in a litre. Trace proteins like this drive both the performance and the cost of a medium (Section 12) (our commentary).

(6) For inducing differentiation

Turning pluripotent stem cells into a specific cell type calls for a different medium. In 2014 Burridge and colleagues reported generating cardiomyocytes from human iPS cells in a chemically defined medium made of just three components: RPMI 1640, L-ascorbic acid 2-phosphate and rice-derived recombinant human albumin, obtaining cell sheets in which up to 95% of cells were positive for a cardiac markerSourced. For neurons, a well-known serum-free combination is “B27-supplemented Neurobasal”, reported by Brewer and colleagues in 1993Sourced. According to the review by Yao and Asayama, B-27 contains ITS plus progesterone, putrescine, triiodothyronine, fatty acids, vitamin E, BSA and glutathioneSourced. It is striking that RPMI-1640, a medium first reported in 1967, became the basal medium for cardiac differentiation in the 2010s (our commentary).

(7) For organoids

Organoid media (organoids being small organ-like structures that self-organise from stem cells) are built to recreate the signals of the body's stem-cell “home”, the niche. In 2009 Toshiro Sato and colleagues reported that a single Lgr5-positive stem cell from mouse small intestine could form an organoid with crypt-villus structuresSourced. According to a review by Clevers and colleagues, this culture used a growth-factor cocktail of EGF, the BMP inhibitor Noggin and R-spondin, within an extracellular matrixSourced. A 2011 report found that long-term culture of human small intestine and colon required the addition of Wnt3A, plus nicotinamide, an Alk inhibitor and a p38 inhibitorSourced. Organoids are covered in more detail in our explainer on organoids and spheroids.

8. The types of culture media at a glance

The table below brings everything so far together along six axes. The same medium can appear under more than one axis (E8, for instance, is both “for pluripotent stem cells” and “chemically defined”).

All Sourced (see the reference list for each row's source). The “Axis” and “In a phrase” classifications, however, are this article's own framing, not an established industry classification.

9. By culture mode: feed media and perfusion media

Even for the same cell and the same goal, the way the medium is supplied changes the medium you need.

Feed media (fed-batch culture)

In fed-batch culture, concentrated nutrients (a feed medium) are added during the run. Kishishita and colleagues reported adding to the feed medium the amino acids that had been heavily consumed in control cultures, and controlling the additions so that waste products such as lactate and ammonia did not accumulate too farSourced. They also added poorly soluble tyrosine in the form of dipeptides and tripeptides to make it dissolve more readilySourced.

Perfusion media (perfusion culture)

In perfusion culture, the cells are kept in the vessel while fresh medium is continuously fed in and spent medium drawn off. In 2006 Konstantinov and colleagues introduced the “equivalent specific perfusion rate” as a common yardstick for comparing fed-batch and perfusion, and showed that the key to making perfusion economically competitive is to cut the dilution rate (how fast the medium is exchanged) sharply while keeping the cell density highSourced. They named this optimisation procedure “push-to-low”, a stepwise lowering of the specific perfusion rate, and reported raising the titre of an antibody against TNF several-foldSourced. A perfusion medium, in other words, has to be concentrated enough to support more cells with less liquid (our commentary).

10. A materials engineer's view (2): adding ingredients that will not dissolve or will not last

11. Format and manufacture: powder, liquid, concentrate, and sterile filtration

Media are supplied as powder, as ready-to-use liquid or as concentrate. The review by Ritacco and colleagues identifies consistency and scalability in preparing powder and liquid media as key issues for industrial useSourced. Powder is light and easy to store, but the user has to dissolve it, adjust the pH and sterilise it. Liquid saves that effort, but it means shipping water, and the stability of the components during storage becomes an issue (our commentary).

12. Media for cultivated meat: the cost wall

Nowhere is the cost of media scrutinised more harshly than in cultivated meat. In a 2021 review, O'Neill and colleagues at UC Davis point out that therapeutic monoclonal antibodies, the largest industrial use of animal cell culture, sell for orders of magnitude more than meat, and that cultivated meat needs a medium that is food-grade, of minimal cost, able to control large-scale proliferation and differentiation, organoleptically acceptable and free of animal componentsSourced.

In 2021 Humbird assessed large-scale production of cultivated meat using the techno-economic methods of industrial fermentation and biopharmaceuticalsSourced. In a scenario of 100,000 tonnes a year (100 kTA), on the scale of the global market, he estimated production costs of US$37 per kg of wet cell mass for fed-batch culture and US$51 for perfusion, of which amino acids account for US$19/kg, growth factors US$3 to 4/kg and glucose about US$0.24/kgSourced. His conclusion is that the economics of such facilities are likely to stand in the way of an affordable food productNot yet confirmed.

Work on bringing costs down is under way. The B8 medium mentioned earlier cut reagent cost to 3% of commercial mediaSourced. In 2022 Stout and colleagues at Tufts University reported that “Beefy-9”, which is simply B8 plus one added component, recombinant albumin, expanded bovine muscle satellite cells over seven passages with an average doubling time of 39 hoursSourced. These are research-stage results, however, and within the scope of this article no record could be confirmed of their use in volume production of foodNot yet confirmed.

13. A materials engineer's view (3): a pharmaceutical formulation at a food price

14. Open problems, and what this article could not confirm

(1) The terms are not consistently defined

“Serum-free”, “animal-component free”, “xeno-free” and “chemically defined” cover subtly different ground depending on the paper or company. Within the scope of this article, no official standard defining them consistently could be confirmed, so the five stages in Fig. 4 are presented as this article's own framing.

(2) The composition of most commercial media is not disclosed

Most commercial media for CHO cells, HEK293 cells, T cells and so on do not have published compositions. This article gives specific components only where the composition could be confirmed in an original paper (E8, the original form of IMDM, the three-component cardiac differentiation medium and so on).

(3) Media costs for cultivated meat are still estimates

Humbird's figures are estimates for a hypothetical scale of 100,000 tonnes a yearNot yet confirmed. Within the scope of this article, no primary source could be confirmed showing what media actually cost in commercial production.

(4) Original papers whose full text could not be checked

For the original papers from the 1950s to the 1970s (Eagle, Dulbecco, Ham, Moore, Stanners, Morgan, Leibovitz), the bibliographic details were checked on PubMed, but the full texts could not be read for this article. Target cells and formulation features follow the description in a peer-reviewed review (Yao and Asayama 2017).

15. Glossary

16. References (primary sources)

1. Eagle H. “Nutrition needs of mammalian cells in tissue culture”, Science 122:501 (1955) — doi.org
2. Eagle H. “Amino acid metabolism in mammalian cell cultures”, Science 130:432 (1959) — doi.org
3. Dulbecco R, Freeman G. “Plaque production by the polyoma virus”, Virology 8:396 (1959) — doi.org
4. Ham RG. “Clonal growth of mammalian cells in a chemically defined, synthetic medium”, PNAS 53:288 (1965) — doi.org
5. Moore GE, Gerner RE, Franklin HA. “Culture of normal human leukocytes”, JAMA 199:519 (1967) (PubMed) — pubmed.ncbi.nlm.nih.gov
6. Stanners CP, Eliceiri GL, Green H. “Two types of ribosome in mouse-hamster hybrid cells”, Nature New Biology 230:52 (1971) — doi.org
7. Iscove NN, Melchers F. “Complete replacement of serum by albumin, transferrin, and soybean lipid in cultures of lipopolysaccharide-reactive B lymphocytes”, J Exp Med 147:923 (1978) — doi.org
8. Barnes D, Sato G. “Methods for growth of cultured cells in serum-free medium”, Anal Biochem 102:255 (1980) — doi.org
9. Yao T, Asayama Y. “Animal-cell culture media: History, characteristics, and current issues”, Reprod Med Biol 16:99 (2017) (PMC) — pmc.ncbi.nlm.nih.gov
10. Morgan JF, Morton HJ, Parker RC. “Nutrition of animal cells in tissue culture; initial studies on a synthetic medium”, Proc Soc Exp Biol Med 73:1 (1950) — doi.org
11. Leibovitz A. “The growth and maintenance of tissue-cell cultures in free gas exchange with the atmosphere”, Am J Hyg 78:173 (1963) — doi.org
12. van der Valk J et al. “Fetal Bovine Serum (FBS): Past – Present – Future”, ALTEX 35:99 (2018) — doi.org
13. European Medicines Agency (EMA) “Guideline on the use of bovine serum in the manufacture of human biological medicinal products”, EMA/CHMP/BWP/457920/2012 rev 1 (PDF) — ema.europa.eu
14. Chen G et al. “Chemically defined conditions for human iPSC derivation and culture”, Nat Methods 8:424 (2011) (PMC) — pmc.ncbi.nlm.nih.gov
15. Ludwig TE et al. “Derivation of human embryonic stem cells in defined conditions”, Nat Biotechnol 24:185 (2006) — doi.org
16. Ludwig TE et al. “Feeder-independent culture of human embryonic stem cells”, Nat Methods 3:637 (2006) — doi.org
17. STEMCELL Technologies “mTeSR™1” product page — stemcell.com
18. Nakagawa M et al. (Kyoto University CiRA and others) “A novel efficient feeder-free culture system for the derivation of human induced pluripotent stem cells”, Sci Rep 4:3594 (2014) — doi.org
19. Ritacco FV, Wu Y, Khetan A. “Cell culture media for recombinant protein expression in Chinese hamster ovary (CHO) cells: History, key components, and optimization strategies”, Biotechnol Prog 34:1407 (2018) — doi.org
20. Kishishita S et al. (Chugai Pharmaceutical and others) “Optimization of chemically defined feed media for monoclonal antibody production in Chinese hamster ovary cells”, J Biosci Bioeng 120:78 (2015) — doi.org
21. Konstantinov K et al. “The ‘push-to-low’ approach for optimization of high-density perfusion cultures of animal cells”, Adv Biochem Eng Biotechnol 101:75 (2006) — doi.org
22. Grieger JC, Soltys SM, Samulski RJ. “Production of recombinant adeno-associated virus vectors using suspension HEK293 cells and continuous harvest of vector from the culture media for GMP FIX and FLT1 clinical vector”, Mol Ther 24:287 (2016) (PMC) — pmc.ncbi.nlm.nih.gov
23. Grace TDC. “Establishment of four strains of cells from insect tissues grown in vitro”, Nature 195:788 (1962) — doi.org
24. Vaughn JL et al. “The establishment of two cell lines from the insect Spodoptera frugiperda”, In Vitro 13:213 (1977) — doi.org
25. Medvec AR et al. “Improved expansion and in vivo function of patient T cells by a serum-free medium”, Mol Ther Methods Clin Dev 8:65 (2018) (PMC) — pmc.ncbi.nlm.nih.gov
26. Burridge PW et al. “Chemically defined generation of human cardiomyocytes”, Nat Methods 11:855 (2014) (PMC) — pmc.ncbi.nlm.nih.gov
27. Brewer GJ et al. “Optimized survival of hippocampal neurons in B27-supplemented Neurobasal, a new serum-free medium combination”, J Neurosci Res 35:567 (1993) — doi.org
28. Sato T et al. “Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche”, Nature 459:262 (2009) — doi.org
29. Boonekamp KE, Dayton TL, Clevers H. “Intestinal organoids as tools for enriching and studying specific and rare cell types”, J Mol Cell Biol 12:562 (2020) (PMC) — pmc.ncbi.nlm.nih.gov
30. Sato T et al. “Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium”, Gastroenterology 141:1762 (2011) — doi.org
31. Berthois Y, Katzenellenbogen JA, Katzenellenbogen BS. “Phenol red in tissue culture media is a weak estrogen”, PNAS 83:2496 (1986) (PMC) — pmc.ncbi.nlm.nih.gov
32. Good NE et al. “Hydrogen ion buffers for biological research”, Biochemistry 5:467 (1966) — doi.org
33. US Food and Drug Administration (FDA) “Guidance for Industry: Sterile Drug Products Produced by Aseptic Processing — Current Good Manufacturing Practice”, September 2004 (PDF) — fda.gov
34. Humbird D. “Scale-up economics for cultured meat”, Biotechnol Bioeng 118:3239 (2021) (PMC) — pmc.ncbi.nlm.nih.gov
35. Kuo HH et al. “Negligible-cost and weekend-free chemically defined human iPSC culture”, Stem Cell Reports 14:256 (2020) (PMC) — pmc.ncbi.nlm.nih.gov
36. Stout AJ et al. “Simple and effective serum-free medium for sustained expansion of bovine satellite cells for cell cultured meat”, Commun Biol 5:466 (2022) (PMC) — pmc.ncbi.nlm.nih.gov
37. O'Neill EN et al. “Considerations for the development of cost-effective cell culture media for cultivated meat production”, Compr Rev Food Sci Food Saf 20:686 (2021) — doi.org

17. Claim-to-source audit

Last updated 23 September 2026. Sources are limited to primary material (original papers and peer-reviewed reviews, published documents from the EMA and FDA, and companies' own product pages). No market-size or market-share estimates are used. Because the article includes structural readings and materials-design interpretations, those are marked as Commentary and kept separate from sourced fact. NK cell media, the specific compositions of commercial media, actual costs in commercial cultivated-meat production, and an official standard defining media terminology are not covered, because they could not be confirmed in published primary sources. All figures are for explanation. Figs. 1, 2, 4, 6 and 7 are concept diagrams; Figs. 3, 5 and 9 are drawings that include our calculation; the hero image and Fig. 8 are AI-generated images, and none of them shows a real product, piece of equipment or medium. This article explains technology. It is not medical advice and does not recommend any product.