Resazurin

Resazurin (Diazoresorcinol) is a water-soluble, non-toxic, stable, membrane-permeable blue non-fluorescent dye (faintly fluorescent). Resazurin is used as a redox indicator, can be reduced to pink, highly fluorescent Resorufin (Ex=530-560 nm, Em=590 nm) in living cells. Resazurin can be used for the detection of cell viability, toxicity, proliferation, migration and invasion in cells (human, plant and animal, bacterial and fungal)^[1][2]. In Vitro:Resazurin (Diazoresorcinol) is commonly used to measure bacterial and eukaryotic cell viability through its reduction to the fluorescent product resorufin. No viable bacteria are detected 24 h post-inoculation following inclusion of Resazurin sodium in TSBc cultures of F. tularensis LVS at the concentration of 44 μM. Lowering the Resazurin concentration to as little as 4.4 μM still results in a 10-fold reduction in viable F. tularensis LVS compare to growth medium alone. Both Resazurin treatments result in a significant decrease in viable F. tularensis LVS bacteria over 22 h. Treatment with Resazurin significantly reduces the number of viable F. tularensis LVS bacteria in HEK293 cells 22 h post-infection^[3].

Guidelines (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs).
1. Solution preparation^[2][3]
1.1 Preparation of stock solution
Solvent: DMSO or ddH2O
Concentration: 1 mg/mL (optimized according to the experiment).
Storage: Store at -20°C or -80°C in dark after aliquoting. Avoid repeated freezing and thawing.
1.2 Preparation of working solution
Dilute to 1 µg/mL with PBS or serum-free medium (optimized according to the experiment).
Note: The working solution should be prepared and used immediately. Keep it away from light.

2. Cell viability
1. Defrost the Resazurin solution in a 37°C water bath.
2. Place the cells on a 96-well plate, and washed by PBS (avoid light).
3. Remove the PBS wash, then add 500 μL (1 μg/mL) of Resazurin solution.
4. Place the plate, in the incubator for 30 min (incubation time depends on cell type and cell number).
5. Measure Resazurin fluorescence (Ex=530-560 nm, Em=590 nm) using a microplate reader.