Cell-based assays are used to screen multiple compounds and determine their viability (cell viability assays), proliferation capacity (cell proliferation assay), and direct cytotoxic effects (cytotoxicity assays). Besides, cell-based assays can measure receptor binding, monitor signal transduction events, track cellular components, and assess organelle function. Regardless of the type of multiplexed ELISA assay used, identifying and determining the population of viable cells remains a critical aspect of bioanalytical experiments.
Today, several cell-based screening assays are used to determine the total number of viable cells at the end of an experiment. These cell-based functional assays offer insights into cell functions and assess the viability or cytotoxicity of chemical compounds. The current article discusses cell-based assays for evaluating cell viability and its importance in drug development.
The significance of cell viability assays in drug development
A cell viability assay considers the ratio between live and dead cells. This assay evaluates the viability of eukaryotic cells in cell culture to analyze cell-mediated cytotoxicity effects and monitor cell proliferation. Several cell viability assay methods are available, including the surface viable count, roller tube technique, fluorogenic dye assay, dilution method, trypan blue, cell viability assay, and nalidixic assay method.
Another central assay method for cell viability assessment is the MTT assay. This method is sensitive and applicable to determine cytotoxicity and cytostatic activity. Cell viability assays can identify drug effects on cells and optimize cell culture conditions. Notably, the factors for defining cell viability can range from cellular enzyme activity, cell membrane integrity, and redox potential of cells.
The protease activity, resazurin reduction, and tetrazolium reduction assay can determine some aspects of enzyme activity or general metabolism as an indicator of viable cells. All these assay formats require incubating the reagent with viable cells to produce a fluorescent or colored product that can be identified using a plate reader. In a standard culture medium, incubating viable cells with substrate will generate an output proportional to the total number of viable cell populations. After cells die, they can’t convert this substrate into a product. This mechanism is employed in all commonly referred cell viability assays. However, the ATP cell viability assay is different. In this assay type, the reagent is not incubated with the viable cells, but adding the reagent ruptures the viable cells immediately.
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Each assay type has its advantages and drawbacks. The ATP detection viability assay is the most sensitive rapid assay with the least degree of interference. On the other hand, the resazurin or tetrazolium reduction assays are less expensive and can deliver adequate assay performance if proper design is followed. Additionally, the fluorogenic cell permeable protease substrate can serve as confirmatory or orthogonal method alternatives, as the substrate is comparatively less cytotoxic than resazurin or tetrazolium compounds.
Conclusion
Cell viability assays are crucial for determining the population of viable cells at the end of an experiment run. Modern cell viability assays incorporating shrimp luciferase can help capture real-time data and offer multiplexing capacities. However, consistent sources, tightly controlled experiments, and thorough characterization of the incubation period and reagent concentration are critical for generating successful and reproducible cell viability assays.
