There is no fixed culture conditions for culturing a certain type of cells. Cells cultured in MEM are likely to grow easily in DMEM or M199. In short, MEM is the first choice for adherent cell culture; RPMI-1640 is a good start for suspension cell and human leukemia cell monolayer culture. It is also widely used in the cultivation of mammalian cells and hybridoma cells, such as human myeloma cells, Mouse hybridoma cells, human leukocytes, B cells and T cells; AIM V (12005) medium (SFM) is the best choice for serum-free culture for various purposes. The medium for selecting cells can also be queried on the ATCC. ATCC (American Type Culture Collection) has collected detailed information on most cells. Open the Cells and hybridomas link on the ATCC webpage and enter the cell name to search the ATCC cell database. The database contains a detailed description of each cell type, including the source of the cells, culture and freezing conditions, and relevant literature.
The same medium will also be used in different cell cultures and different experimental needs due to different additives. The functions of various additives in the medium will be described in detail below.
1. Is L-Glutamine important in cell culture? Is it unstable in solution?
It is an essential amino acid for cell growth and provides an important source of energy for cultured cells. After removing the amino group, L-glutamine can be used as an energy source for culturing cells, participating in protein synthesis and nucleic acid metabolism. L-Glutamine will degrade in the solution after a period of time, and the degradation rate changes with the storage temperature. The degradation of L-glutamine leads to the formation of ammonia, which is toxic to some cells.
2. What is GlutaMAX-I? How to use GlutaMAX-I in cultured cells? How stable is this dipeptide?
GlutaMAX-I, which is glutamic acid dipeptide, is a derivative of L-glutamine, and its unstable alpha-amino group is protected with L-alanine. A peptidase gradually cleaves dipeptides, releasing L-glutamine for use. GlutaMAX-I dipeptide is very stable. Even when sterilized at 121 pounds for 20 minutes, the GlutaMAX-I dipeptide solution has minimal degradation. If under the same conditions, L-glutamine is almost completely degraded.
3. What is the role of sodium pyruvate in the medium?
Sodium pyruvate can be used as an alternative carbon source in cell culture. Although cells prefer glucose as a carbon source, cells can also metabolize sodium pyruvate if there is no glucose.
4. What are the essential functional differences between Hank's Balanced Salt Solution (HBS) and Earle's Balanced Salt Solution (EBS)?
The main difference between HBS and EBS is the level of sodium bicarbonate, which is higher in Eagle's (2.2g / L) than in Hanks' (0.35g / L). Sodium bicarbonate needs to be balanced with high levels of CO2 to maintain the pH of the solution. Eagle's solution will become alkaline in CO2 at air level, and Hanks' solution will become acidic in CO2 incubator. If you want to save the tissue in a CO2 incubator, you need to use Eagle's solution. If only the tissues to be stored in the cell culture medium are washed, Hanks' solution is sufficient.
5. How does the pH of the culture medium affect cell growth?
Since most cells have a pH of 7.2-7.4, deviation from this range may have a detrimental effect on cell growth. However, the pH requirements of various cells are not exactly the same. Primary cultured cells generally have poor tolerance to pH changes, and infinite cell lines have strong tolerance. But overall, cells are more resistant to acid than alkaline. When preparing the culture solution, it is necessary to pay attention to the following: when the newly prepared medium is filtered through a 0.10um or 0.22um filter membrane, the pH of the solution will also float upward by about 0.2.
6. Should I use 5% or 10% CO2 when culturing cells, or does it have no effect at all?
Answer: Most of the medium uses HCO3- / CO32- / H + as the pH buffer system, and the NaHCO3 content in the medium will determine the CO2 concentration that should be used in cell culture. When the NaHCO3 content in the medium is 3.7 g per liter, 10% CO2 should be used for cell culture; when the NaHCO3 in the medium is 1.5 g per liter, 5% CO2 should be used to culture the cells.
7. The role of Hepes
HEPES solution: It is a weak acid, the Chinese name is hydroxyethyl piperazine ethanesulfonic acid, the main role is to prevent the pH of the medium from changing rapidly. Under open culture conditions, when observing the cells, the culture medium deviated from the 5% CO2 environment, the CO2 gas quickly escaped, and the pH increased rapidly. If HEPES is added, the pH can be maintained at around 7.0 at this time. Generally, HEPES should be added when performing clonal culture. Can be used during the primary culture of nerve cells
8. NaHCO3
The NaHCO3-CO2 buffer system is used in the culture medium, and the open culture is used to make the CO2 generated by cell metabolism overflow the culture bottle in time, and then the CO2 concentration (5%) in the thermostat is stably adjusted to be in equilibrium with the NaHCO3 in the culture medium , Thereby adjusting the PH value of the culture medium.
9. NEAA: non-essential amino acids
NEAA (non-essential amino acids) is a mixture containing several non-essential amino acids. The original medium added to it is different. NEAA also has different types. For example, NEAA added to MEM contains L-Alanine, L-Asparagine, L-Aspartic Acid , L-Glutamic Acid, L-Glycine, L-Proline, L-Serine .
10. Calcium chloride
Calcium chloride is toxic to osteoblast culture, so the culture medium of osteoblasts should not contain calcium chloride.
11. What medium can omit phenol red?
Phenol red is used as a pH indicator in the medium: red when neutral, yellow when acidic, and purple when alkaline. Phenol red itself does not affect the quality of biological products and can be removed by purification techniques, but phenol red may cause intracellular sodium / potassium imbalance in serum-free medium and affect cell growth. Of course, this effect can be affected by serum And or lighten. Phenol red is not an essential component in the culture medium. Many foreign vaccine or antibody manufacturers use phenol red-free medium in the production process.
Studies have shown that phenol red can mimic the effects of steroid hormones (especially estrogen). To avoid sterol reactions, use phenol red-free medium when culturing cells, especially mammalian cells. Because phenol red interferes with the detection, some researchers do not use phenol red-added medium when doing flow cytometry.
The chemical formula of this product is C3H7NO3, named after the silk from which it was first derived.Serine is a neutral aliphatic hydroxyl-containing amino acid, a non-essential amino acid.Serine plays a role in the metabolism of fats and fatty acids and in the growth of muscle, in the manufacture and processing of cell membranes, and in the synthesis of muscle tissues and of sheaths enclosing nerve cells.D-Serine is an essential amino acid that is used in the manufacture of cell membranes and in the synthesis of muscle tissues and sheaths enclosing nerve cells. It is mainly used in compounded amino acid preparations for amino acid supplementation.D-serine is also an important neurotransmitter, and the N-methyl-D-aspartate (NMDA) receptor is an important endogenous ligand for it, both of which play important roles in the central system. Production methods usually include fermentation, protein hydrolysis and extraction, chemical synthesis, and biological enzymes.
D-serine,D-Ser-OH,312-84-5,C3H7NO3
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