Cell viability is a fundamental measurement in biological research, representing the proportion of healthy, live cells within a given population. This metric provides insights into cellular health and function under specific conditions. Understanding cell viability is important for various scientific investigations, from basic cellular biology to advanced medical applications.
What Cell Viability Means
Cell viability quantifies the percentage of cells in a sample that are alive and functioning. Live cells maintain intact cell membranes and active metabolic processes, allowing them to regulate their internal environment. In contrast, dead cells often have compromised membranes and cease metabolic activity. Cells can also undergo programmed cell death (apoptosis) or uncontrolled cell death (necrosis), both of which impact the overall viable cell count.
The Cell Viability Calculation
The cell viability formula is straightforward, expressed as a percentage. It is calculated by dividing the number of viable cells by the total number of cells in a sample, then multiplying by 100%.
For instance, if a researcher counts 80 live cells in a sample containing 100 total cells (including both live and dead), the calculation would be (80 / 100) x 100% = 80%. The “number of viable cells” refers to those exhibiting characteristics of life, while “total number of cells” includes all cells present, regardless of their state. This ratio allows for standardized comparisons across different experiments.
Techniques for Measuring Viability
Accurately determining the number of viable and total cells for the viability formula requires specific laboratory techniques.
Trypan Blue Exclusion Assay
This widely used method relies on the integrity of the cell membrane. Live cells with intact membranes exclude the Trypan blue dye, appearing clear under a microscope. Dead cells with compromised membranes allow the dye to enter, staining their cytoplasm blue. Researchers mix a cell suspension with the dye and then manually count clear and blue cells using a hemocytometer.
Metabolic Assays
Metabolic assays, such as MTT, XTT, and Resazurin (AlamarBlue), measure the metabolic activity of live cells. These assays involve the conversion of a soluble, often colorless, substrate into a detectable, colored product by enzymes present in metabolically active cells, particularly mitochondrial dehydrogenases. For example, in the MTT assay, yellow MTT tetrazolium salt is reduced to purple formazan crystals by these enzymes, with the color intensity directly proportional to the number of viable cells. These methods are employed for high-throughput screening due to their quantitative nature.
ATP Assays
ATP assays quantify adenosine triphosphate (ATP) levels within cells, which serve as a marker for metabolically active cells. Living cells continuously produce ATP, and a decrease in ATP levels indicates cell stress or death. These assays commonly use the firefly luciferase enzyme, which reacts with ATP to produce light, with the luminescence intensity directly correlating to the amount of ATP and, consequently, the number of viable cells. This bioluminescent approach offers high sensitivity and a broad dynamic range.
Fluorescent Stains
Fluorescent stains, such as Calcein AM and Ethidium Homodimer-1 (EthD-1), offer another approach to differentiate live and dead cells. Calcein AM is a cell-permeant, non-fluorescent dye that is converted into a green fluorescent product by intracellular esterases in live cells, which is then retained within the cell. Conversely, EthD-1 is a red fluorescent dye that can only enter cells with damaged or compromised membranes, binding to nucleic acids in dead cells. This two-color staining allows for simultaneous visualization of live and dead cells under a fluorescence microscope or can be quantified using flow cytometry, providing detailed information on membrane integrity and enzymatic activity.
Applications of Cell Viability
Cell viability measurements are extensively applied across various scientific and medical fields, providing insights into cellular responses to different conditions.
Drug Discovery and Development
Cell viability assays are used to assess the toxicity and efficacy of new pharmaceutical compounds. Researchers can determine if a drug candidate kills target cells (cytotoxicity) or merely inhibits their growth (cytostasis), helping to identify promising therapeutic agents and establish safe dosage ranges.
Toxicology Studies
Toxicology studies rely on cell viability assays to evaluate the harmful effects of chemicals and environmental contaminants on living cells. These assessments help determine safe exposure levels for various substances, supporting regulatory assessments and public health protection. By observing changes in cell viability, scientists can identify potential hazards early in the research process.
Cell Culture Maintenance
Monitoring cell viability is a standard practice to ensure the health and quality of cell lines used in ongoing research. Regular viability checks help identify issues such as contamination, nutrient depletion, or suboptimal growth conditions, allowing researchers to adjust protocols to maintain healthy cell populations.
Tissue Engineering and Regenerative Medicine
Tissue engineering and regenerative medicine also depend on accurate cell viability assessments. These measurements are used to evaluate the survival and functionality of cells incorporated into engineered tissues or those prepared for transplantation. Ensuring high cell viability is important for the successful integration and performance of these biological constructs in therapeutic applications.
Basic Biological Research
Cell viability assays are important in basic biological research. They help scientists understand how cells respond to diverse environmental changes, such as stress, nutrient availability, or changes in temperature or pH. This foundational knowledge contributes to a broader understanding of cellular processes and disease mechanisms.