Cell proliferation is one of the important physiological functions of living cells and is an important life characteristic of organisms. Cell proliferation is the basis of organism growth, development, reproduction as well as heredity.
Normal cells are metabolically vigorous and their mitochondria contain succinate dehydrogenase, which can reduce tetrazolium salts (e.g., MTT, XTT, WST-1, and WST-8, etc.) to purple crystalline substances that deposit around the cell. Then we can use a microplate reader to read the OD value (optical density), thus detecting the status of cell proliferation.
Cell counting kit-8 (also known as CCK-8) is a rapid and highly sensitive assay based on WST-8 (Water Soluble Tetrazolium-8) and widely used for cell proliferation analysis.
The mechanism is: WST-8 [chemical name: 2-(2-Methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium Sodium Salt] is reduced by dehydrogenases in cells under the reaction of the electron carrier 1-methoxy-5-methylphenazinium sulfate dimethyl ester (1-methoxy PMS) to a highly water-soluble yellow formazan product (formazan dye).The number of formazans generated directly correlates to the number of living cells.This property can therefore be exploited to directly carry out cell proliferation assays.The more rapidly the cells proliferate and the more cells proliferate, the darker the color is. For the same cells, there is a linear relationship between the intensity of the color and the number of cells.
Deng, Danni et al. “p62 acts as an oncogene and is targeted by miR-124-3p in glioma.” Cancer cell international vol. 19 280. 6 Nov. 2019, doi:10.1186/s12935-019-1004-x
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5~10 business days, depending on cell growth and experimental design
cell lines, and experimental conditions parameter settings
raw data, analytical results, experimental reports
Other methods to detect cell proliferation
|BrdU can be inserted into replicated DNA duplexes in place of thymidine, and an immunological assay is used to determine the amount of BrdU in DNA and thus detecting the proliferative capacity of cells
|Direct determination of DNA synthesis; can be combined with other markers, double staining, identify the types of proliferating cells
|Ki-67 is mainly expressed in the G1, s, G2 and M phases of cell division, but is not expressed in the G0 phase. Therefore Ki-67 can be used to measure the growth index of cells
|Imaging results can intuitively reflect the spatial distribution of proliferating cells; the stimulatory effect or antiproliferative effect of drugs and so on can be obtained
|Viable cells can be counted for their ability to detect proliferation by using trypan blue that is unable to permeate through intact membranes of viable cells thus no staining but is permeable to dead cells
|Simple assays, and cells can be directly counted. Not suitable for assessing specific subpopulations and a larger number of cells
|Colony formation rate, i.e., survival rate of cell seeding, represents adherent survival of the seeded cells and the number of clones formed, reflecting the population dependence and proliferative capacity of the cells
|Suitable for cells that are adherent and cells with anchorage independent growth