Resazurinbased

Resazurin-based assays are a class of biochemical tests that utilize resazurin, a blue, water-soluble dye, as a redox indicator. In viable cells, resazurin is reduced by cellular metabolic activity to resorufin, which is pink and highly fluorescent, and further to dihydroresorufin, which is colorless. The extent of dye reduction correlates with metabolic rate, enabling quantitative assessments of viability, proliferation, cytotoxicity, or microbial growth.

Resazurin-based readouts are widely used in cell biology and microbiology. In mammalian cell assays, products such as Alamar Blue and CellTiter-Blue are employed to monitor viability in adherent or suspension cultures. In microbiology, resazurin reductions indicate microbial respiration in diluted samples or growth in colorimetric assays and can be used in environmental monitoring and antimicrobial testing.

Readout can be colorimetric, measuring absorbance at around 590–600 nm for the blue oxidized form, or fluorometric, taking advantage of resorufin fluorescence with excitation near 560–570 nm and emission near 585–590 nm. Many protocols add a defined volume of resazurin solution to wells containing cells or samples and incubate for a period ranging from a few hours to overnight, depending on cell type and assay sensitivity.

Advantages include simplicity, low toxicity at assay-relevant concentrations, and compatibility with high-throughput formats. Considerations include potential interference from strong reducing media or serum supplements, nonlinear response at high cell numbers, and the need for appropriate controls and standard curves. Proper storage and protection from light prolong dye stability.