Cell Viability Meaning

Cell viability is the measurement of the number of live cells in a sample, typically given as a proportion of the total cell population. It is a critical parameter to measure when assessing the health of any cell culture or outcome of an assay (e.g., transfection).

Cell count assays are applicable for every cell type and culture condition, including cell suspensions and adherent cells. During initial seeding or passaging, it is important to plate the correct number of live cells per area or volume for adherent cultures or bioreactors. Large numbers of dead cells can be detrimental to growth as they release proteins and other cellular factors like cytokines that negatively affect viable cells.

Cell viability assays represent a crucial testing component during the drug development process. These assays can inform on the effectiveness and toxicity of medications, such as anticancer medications, where cell survival is monitored.

Common Cell Viability Assays

• Trypan blue exclusion assay : This assay identifies viable cells based on the exclusion or uptake of Trypan blue. Viable cells do not permit the absorption of the dye, while dead cells will be colored blue due to the lack of intact cell walls. The assay can be performed manually or automated in most pharmaceutical labs. A traditional light microscope is used for assay readout.
• Propidium iodide staining : Propidium iodide (PI) assay is a commonly used flow cytometry assay. Similar to Trypan blue, PI only crosses the membrane of dead or dying cells. Once inside the cell, it binds to the DNA found in the nucleus, resulting in the emission of red fluorescence. The flow cytometer quantifies the number of viable cells at the single cell level. PI is often combined with other cellular markers to provide clues about viable cellular subpopulations, especially when researching targeted therapies against specific cell populations in a mixture.
• MTT assay : The MTT viability assay is a commonly used colorimetric method to evaluate the metabolic activity of cells and can serve as a real-time viability assay. The MTT reagent can penetrate both the cell membrane and the inner mitochondrial membrane of viable cells. Cells with active metabolism reduce the MTT reagent to formazan, resulting in a measurable color change. The MTT assay requires a longer incubation time and a spectrophotometer for the final readout.
• ATP assay : Adenosine triphosphate (ATP) is produced by living cells and used as a readout to assess cell viability. The most common variation of the assay uses the bioluminescence from a firefly luciferase resulting from interaction with viable cellular ATP. ATP assays are very popular within pharmaceutical labs performing high-throughput screening workflows because of their quick turnaround time.

Factors Affecting Cell Viability

• When conducting viability assays for cells in culture, it is important to carefully control and maintain the factors influencing viability like excessive handling or composition of cell culture medium. Cells require an optimal temperature range for proper functioning, and deviations from this range can induce cellular stress and result in cell death. Similarly, maintaining the appropriate pH is crucial. Any deviations from physiologically optimal ranges can disrupt cellular processes and eventually lead to cell death. For instance, acidic or alkaline conditions can interfere with enzyme activity and disrupt protein structure, leading to cellular dysfunction.
• Cell viability is also heavily influenced by the availability of nutrients and oxygen. Adequate nutrient supply, including essential amino acids, glucose, growth factors and vitamins, is vital for supporting cellular metabolic activities. Insufficient availability of these nutrients can induce cellular stress and ultimately lead to cell death. Cells require oxygen for efficient energy production through aerobic respiration. When oxygen levels are too low (hypoxia), cellular stress occurs, and cells may undergo cell death. Conversely, excessive oxygen levels (hyperoxia) can result in oxidative stress, causing damage to cellular structures.
• Cellular stressors, including oxidative stress, heat shock, and DNA damage, can significantly affect cell viability. Oxidative stress occurs when there's an imbalance between ROS production and antioxidant defense mechanisms. Heat shock, characterized by sudden exposure to high temperatures, can disrupt the proper folding of proteins, resulting in cellular dysfunction and death. DNA damage caused by radiation or chemical exposure can activate cell death pathways.
• Cell viability can be influenced by environmental factors such as radiation, chemical exposure and pathogens. Ionizing radiation, for instance, can induce DNA damage and disturb cellular processes, potentially compromising cell viability. Exposure to toxic chemicals can result in cellular stress and eventual cell death. Pathogens, including bacteria and viruses, can invade cells and disrupt their normal functions, leading to cell death .

Strategies for Improving Cell Viability

Maintaining optimal culture conditions and utilizing cell viability, cell toxicity and cell cycle assays are essential for assessing and enhancing cell viability in various research settings. Optimizing factors such as temperature, pH, nutrient availability and oxygen levels are crucial for maintaining cell viability in cell culture. Proper culture conditions provide cells with the necessary resources for growth and survival. Supplementing the culture medium with cell viability-enhancing compounds like growth factors and antiapoptotic molecules can support cell growth and improve viability. Protective agents can shield cells from harsh conditions and prevent cell death, while antioxidants can mitigate oxidative stress. Additionally, genetic modifications can be employed to enhance cell survival. For example, inhibiting specific genes has been shown to improve cell viability and enhance the expression of key survival genes in certain cancer cells.

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