1. Seed 50 µl (5000 cells) per well in 96 well plate (clear, flat bottom).

2. You may need to add dilution of drugs or other agents at this time or after cells have attached overnight.

3. Incubate cells for the desired time at 37 °C.

4. Add 15 µl of the Dye Solution to each well.

5. Return the plate to the incubator for 4 hours.

6. Add 100 µl of the Solublization/Stop Solution.

7. Let stand for 1 hour at room temperature, mix.

Alternatively let stand for overnight at room temperature in a humidified container.

8. Read the absorbance at 570 nm with a plate reader.