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Cell culture media

Cell culture media

The cell culture media is a nutrient broth that contains growth factors, vitamins, minerals, salts, energy sources, and amino acids cells need to survive, grow and perform cellular functions.

Cell culture medium is a liquid or gel that supports the survival, growth and cellular functions. Animal cells can be cultured in media from natural animal sources such as tissue extracts and biological fluids like plasma, serum and amniotic fluid . Synthetic or artificial media contain nutrients, vitamins, salts, oxygen and carbon dioxide gas phases, blood serum proteins, carbohydrates and cofactors, which are used by enzymes .

Serum containing media

Synthetic media can also contain animal-derived supplements. For example, fetal bovine serum (FBS) is commonly used as a low-cost supplement that provides hormones, growth factors, protective protease inhibitors, chelators or carriers for nutrients, essential nutrients, components that promote cell adherence . Serum also serves to bind and neutralize toxins .

Bovine calf serum, newborn calf serum, fetal bovine serum, horse serum and human serum are used in cell culture.

Serum-free media

Serum-free media is also referred to as ‘defined culture media’. While the composition of serum varies from batch to batch, serum-free media have defined quantities of growth factors, lipoproteins and other proteins normally found in serum . Serum-free media are usually formulated for specific cell types.

The use of serum for cell therapies and therapeutic products produced in cell culture can be a point of entry for pathogens to contaminate products . Clean meat companies up until 2018 have used animal derived serum to culture meat but weaning off of serum is desirable so that aside from the starting cells, the meat products would not rely on animals .

Chemically defined media

These media contain pure protein additives like growth factors which are produced by bacteria or yeast through genetic engineering and also contain vitamins, cholesterol, amino acids and fatty acids . Every ingredient is defined and reproducible, making it easier to identify cellular secretions .

Most stem cells are cultured in serum-free media with specific growth factors and additives which either promote stem cell self-renewal or promote differentiation into specialized cell types . The factor bFGF (basic fibroblast growth factor) is added to regulate stem cell-renewal for human embryonic stem cells (ESCs), induced pluripotent stem cells and neural stem cells . Leukemia Inhibitory Factor (LIF) inhibits differentiation and promotes the expansion of mouse but not human ESCs . Retinoic acid, ascorbic acid, hormones and DNA demethylating agents or intracellular cAMP elevating IBMX can induce stem cells to differentiate down defined pathways .

Protein-free media

Protein-free media such as minimal essential medium (MEM), RPMI-1640, make it easier to purify protein products expressed by the cells and it also provides better cell growth and protein expression . The proprietary Protein free medium (PFM) is a serum-free and protein-free culture medium without hormones or growth factors optimal for culturing Chinese Hamster Ovary (CHO) cells which are used to produce therapeutic proteins .

Basic components

Cell culture media have buffering systems to regulate pH. Natural buffering systems use gaseous CO2 to balance with CO3/HCO3 in the culture medium. Cultures using this system are incubated with an air atmosphere of 5-10% CO2 . HEPES is a buffering agent that does not need a controlled gaseous atmosphere but it is expensive and toxic to some cell types at high concentrations . Commercially available culture media usually contain phenol red as a pH indicator that changes color when pH is not optimal, but it is not suitable for all cell culture applications.

Amino acids are included in media because they are the building blocks of proteins and essential all amino acids cannot be synthesized by the cells and cell proliferation capacity depends on the concentration of amino acids . Nonessential amino acids stimulate growth and extend viability of cells.

Carbohydrates like glucose and galactose provide energy for cells . Proteins like albumin, transferrin and fibronectin are more important additives for serum-free media. Albumen binds water, salts, fatty acids, hormones, vitamins and toxic substances and transports them . Fibronectin is helps cells attach and transferrin transports iron to the cell membrane.

Salts are important for osmotic balance, regulating membrane potential and providing sodium, potassium and calcium ions. Fatty acids, vitamins and trace elements like copper, zinc and selenium are also needed for biological processes . Aprotinin inhibits serine proteases like trypsin, which degrade proteins .

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Animal‐cell culture media: History, characteristics, and current issues

Tatsuma Yao and Yuta Asayama

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