Cell Culture Medium

Cell culture medium is an artificially prepared mixed solution that provides necessary nutrients and suitable environmental conditions for cell growth, reproduction, and metabolism.

Main Components and Their Functions

1. Nutrients
   • Amino Acids: They are the basic units of protein synthesis. Cells cannot synthesize all amino acids by themselves, so the medium needs to provide essential amino acids such as valine, leucine, isoleucine, etc. These amino acids are used to build various proteins in cells and are crucial for maintaining cell structure and function. For example, in muscle cell culture, an adequate supply of amino acids helps maintain the synthesis of actin and myosin in cells, thereby ensuring the contractile function of cells.
   • Carbohydrates: They mainly provide energy for cells. Glucose is the most commonly used carbohydrate. Cells break down glucose through metabolic pathways such as glycolysis and aerobic respiration to produce ATP, which provides power for various life activities of cells. In some special cell cultures, such as tumor cell culture, due to the high metabolic rate of tumor cells, a higher concentration of glucose may be required to meet their energy demands.
   • Vitamins: They play multiple key roles in cell metabolism. For example, vitamin B complex (such as vitamin B1, B2, B6, etc.) is involved in energy metabolism, substance synthesis, and redox reactions in cells. Vitamin C, as an antioxidant, can protect cells from free radical damage. In nerve cell culture, an adequate supply of vitamins helps maintain the normal physiological functions of nerve cells, such as the synthesis of neurotransmitters.
   • Inorganic Salts: They include various ions such as sodium (Na⁺), potassium (K⁺), calcium (Ca²⁺), magnesium (Mg²⁺), and phosphorus (P). These ions play important roles in maintaining osmotic pressure balance, regulating cell membrane potential, and serving as cofactors for enzymes. For example, Ca²⁺ plays a key role in cell signal transduction and can activate multiple signal pathways to participate in cell activities such as proliferation, differentiation, and secretion.
2. Growth Factors
   • They are proteins or polypeptides that can stimulate cell growth, proliferation, and differentiation. For example, epidermal growth factor (EGF) can promote the proliferation and migration of epithelial cells and is widely used in skin tissue engineering and cell culture. Fibroblast growth factor (FGF) is important for the growth and differentiation of many cell types such as fibroblasts and neural stem cells. These growth factors bind to receptors on the cell surface and activate intracellular signal transduction pathways to regulate cell physiological activities.
3. Buffering Systems
   • Cells require a stable pH environment for growth. Commonly used buffering systems are the bicarbonate buffering system (such as NaHCO₃ – H₂CO₃) and the HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) buffering system. During cell culture, cell metabolism produces acidic substances such as lactic acid, and these buffering systems can neutralize acidic substances to maintain the pH of the medium within an appropriate range (generally 7.2 – 7.4). For example, in mammalian cell culture, when cell metabolism is vigorous, the produced lactic acid will cause the pH of the medium to decrease, and the buffering system will play a role to prevent excessive pH reduction from damaging the cells.
4. Other Components
   • Serum: It is an important additive component in cell culture medium. Serum contains a variety of proteins, growth factors, hormones, and nutrients, which can provide rich nutritional support and growth-promoting signals for cells. For example, fetal bovine serum (FBS) contains a large amount of albumin, transferrin, and other nutrients, as well as multiple growth factors such as platelet-derived growth factor (PDGF). However, due to the complex composition of serum and batch-to-batch differences, it may have a certain impact on the reproducibility of experimental results.
   • Antibiotics and Antifungal Agents: To prevent microbial contamination during cell culture, antibiotics (such as penicillin, streptomycin) and antifungal agents (such as amphotericin B) are usually added to the medium. These drugs can inhibit the growth of bacteria and fungi, but it should be noted to control the dosage during use to avoid toxic effects on cells.

Classification and Application Scenarios

1. Natural Media
   • Serum Media: Serum is the main component, such as the aforementioned fetal bovine serum medium. It is rich in nutrients and can support the growth and proliferation of many cell types, and is widely used in basic cell biology research, vaccine production, and other fields. For example, in the production process of virus vaccines, using serum medium to culture host cells can provide good conditions for virus replication.
   • Tissue Extract Media: Prepared by using extracts of animal tissues (such as embryonic tissue, liver tissue, etc.). This medium contains growth factors and nutrients specific to the tissue and is suitable for culturing some cells that are highly dependent on specific tissue signals. For example, in neural stem cell culture, using brain tissue extract medium can provide the special nutrients and signal molecules required for nerve cell growth.
2. Synthetic Media
   • Basic Synthetic Media: Precisely formulated according to the basic nutrients required for cell growth. For example, DMEM (Dulbecco’s Modified Eagle Medium) medium contains various amino acids, vitamins, carbohydrates, etc., which can meet the basic growth requirements of most mammalian cells. This medium is widely used in cell biology research, genetic engineering, and other fields, and is often used to culture cell lines such as HeLa cells (human cervical cancer cell line).
   • Serum-Free Synthetic Media: Does not contain serum components, and replaces the function of serum by adding specific growth factors, proteins, and nutrients. The advantage of this medium is that its composition is clear, which is convenient for analyzing and controlling experimental results, and reduces the batch differences and potential pathogen contamination risks brought by serum. It is increasingly widely used in fields with high requirements for product quality and safety, such as biopharmaceutical production and cell therapy. For example, in the production process of monoclonal antibodies, using serum-free synthetic medium can better control product quality.

Precautions for Use

1. Storage Conditions
   • Cell culture media usually need to be stored at low temperature (2 – 8°C) and protected from light. This is because some components in the medium (such as vitamins, growth factors, etc.) are sensitive to temperature and light, and are prone to decomposition or inactivation under high temperature or light. At the same time, it should be noted to avoid repeated freezing and thawing of the medium, as this may lead to precipitation and deterioration of components.
2. Aseptic Operation
   • When using cell culture media, strict aseptic operation principles must be followed. Because the medium is rich in nutrients, it is very susceptible to contamination by bacteria, fungi, and mycoplasma. During the preparation and use of the medium, sterile containers, utensils, and reagents should be used, and operations should be carried out in a sterile environment such as a laminar flow hood. For example, when adding the medium to the cell culture container, it is necessary to ensure that pipettes, culture flasks, etc. have been strictly sterilized.
3. Quality Control
   • Pay attention to the quality of the medium. When purchasing the medium, choose a reliable supplier, and carefully check the appearance (such as color, transparency, etc.) and expiration date of the medium before use. If the medium shows abnormal phenomena such as turbidity, discoloration, or precipitation, it may indicate that the medium has deteriorated and should not be used. At the same time, for some critical cell culture experiments, it is best to conduct quality tests on the medium, such as detecting the content of nutrients, pH value, etc.
4. Selecting Appropriate Media According to Cell Types
   • Different cell types have different requirements for the composition and environment of the medium. For example, adherent cells (such as fibroblasts) and suspension cells (such as lymphocytes) require different types of media for support. Adherent cells need an appropriate attachment substrate in the medium, and the composition of the medium should be conducive to cell adhesion and growth; suspension cells require the medium to provide sufficient nutrients and space to maintain the suspension state and normal growth of the cells. Therefore, before cell culture, it is necessary to fully understand the characteristics of the cells and select an appropriate medium.
5. Use of Additives
   • When adding serum, growth factors, and other additives to the medium, pay attention to controlling the addition amount and method. For example, the addition amount of serum is generally between 5% and 20%, and too high or too low may affect cell growth. At the same time, when adding additives, they should be slowly and evenly added to the medium to avoid local high concentrations that may damage the cells. For some temperature-sensitive additives, such as growth factors, they should be added at an appropriate temperature to ensure their activity.