The following is the automation protocol developed by
1. Grow Salmonella tester strains in broth at 37 °C overnight with shaking.
2. Prepare test chemical by incubating in rat liver S9 extract.
3. Test chemical is added to the Salmonella culture at different concentrations in a 24-well plate.
4. Incubate at 37 °C for 90 minutes without shaking.
5. Add Salmonella Reversion Indicator Medium (bromocresol purple in Vogel Bonner medium supplemented with amino acids)
to each well. Bromocresol purple is a pH-sensitive dye, which turns color from purple to yellow as wells containing growing (revertant) Ames II colonies reach a pH of 5.2 and lower.
6. Transfer cells in the 24-well plates to the 384-well plates.
Incubate at 37 °C for 2 days.
7. Score for positive (yellow) wells with a plate reader at 492 nm.