Cell Viability Assay

Cell viability is a measurement for evaluating the health and ability of a survival cell population, while at the same time, the evaluation can also indirectly determine the cell toxicity (decrease in cell viability). Typically, measuring cell viability can be used to identify the proportion of healthy cells in a population, optimize culture conditions, or assess the toxicity and efficacy of potential drug treatment. In general, cell viability assays can be roughly divided into several assays with different strategy, including DNA synthesis proliferation assays, cell proliferation staining assays, and metabolic proliferation assays.
DNA synthesis proliferation assay
This cell viability assays employ nucleoside analogues (5-bromo-2’-deoxyuridine, BrdU; 5-ethynyl-2’-deoxyuridine, EdU) that are incorporated into newly synthesized DNA of proliferative cells. The more proliferative cells, the more BrdU or EdU are found in the culture. In BrdU assay, DNA with BrdU can be recognized by anti-BrdU antibody with HRP conjugation. However, in EdU assay, DNA with EdU can be easily labeled by using copper-catalyzed azide-alkyne “click” chemistry to attach fluorescent probes for emission detection. Moreover, there are anti-BrdU with HRP conjugation, which is also used to recognized the incorporated BrdU cells.
Cell proliferation staining assay
Immunohistochemistry (IHC) assay is commonly used to determine cell proliferation. Ki-67, proliferating cell nuclear antigen (PCNA), and minichromosome maintenance (MCM) proteins are well-known cell proliferation marker proteins for recognition. Anti-Ki-67, anti-PCNA and anti-MCM are widely used to analysis the distribution and expression pattern of their specific target in disease versus healthy tissue samples, which are particularly relevant in identifying the effect of treatments on tumor cell proliferation.
Metabolic proliferation assays
WST-8 (5-(2,4-disulfophenyl)-3-(2-methoxy-4-nitrophenyl)-2-(4-nitrophenyl)-2H-tetrazolium) and ATP luciferase assays are two sensitive and convenient cell viability assays for high through put screening. In WST-8 assays, the water soluble and stable tetrazolium salt WST-8 is reduced by cellular dehydrogenases to a soluble orange formazan on the cell surface. The amount of colored formazan produced is directly proportional to the number of living cells and is measured by absorbance at 460 nm. The excellent stability and little cytotoxicity of the WST-8 solution is useful for assays that require long incubation (such as 24 to 48 hours). In addition, ATP luciferase assay is by measuring intracellular ATP levels. Upon cell lysis, the released ATP is used as energy for ATP-dependent luciferase reaction that emits luminescence. The detected light is directly proportional to the amount of lysed cell, refer to cell proliferation. Both of the shift in color of WST-8 assay and the emission light of ATP luciferase can be measured by spectrophotometer and reports on the metabolic activity of cells. These are often employed for high-throughput cell viability measurements.

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