An Overview of Cytotoxicity Assay During Drug Development Cytotoxicity, as the name suggests, is the toxicity of the cells. When the cells are treated with cytotoxic compounds, it may lead to cell necrosis - leading to loss of membrane integrity, and the result is rapid death caused by cell lysis. These cells can stop growing and dividing, or the cells can activate a genetic program leading to controlled death. The absorption of harmful chemicals causes cytotoxicity – the chemicals can kill cancer cells but have the potential to destroy the healthy cells also. In drug development, studying cytotoxicity is necessary because cytotoxic drugs are necessary for treating critical diseases. The drugs contain chemicals which are toxic to cells, preventing their replication or growth and are effective to control diseases such as cancer. The basis of a Cytotoxicity assay assesses the damage to cellular membranes. It follows four types of methods: ● ● ● ●
Enzyme Leakage assays Membrane impermeable dyes Amine-reactive dyes (used in live: dead cell assays) Dye combination live: dead cell assays
Enzyme Leakage: These assays measure the activity of enzymes that leak into the extracellular level causing cell membrane damage. The most popular assay is for lactate dehydrogenase. Membrane Impermeable Dyes: Cell viability assays often use membrane-impermeable fluorescent dyes (mostly DNA stains) that stain cells with damaged cell membranes. DRAQ7 and 7-AAD have largely replaced propidium iodide for cell viability assays due to its broad emission spectra and tendency to bind to live cells. Amine Reactive Dyes for Live: Dead Cell Assays- Amine-reactive dyes weakly stain viable cells by binding to cell surface amines and strongly stain membrane-compromised cells by reacting with intracellular amines. The fluorescence level can differentiate dead and live cells.