H₂O₂ Kill Assays of Planktonic Stationary Phase Bacteria

Materials and Reagents

1. Phosphate buffered saline (PBS) solution (Sigma-Aldrich, catalog number: P4417-100TAB )
   
2. 30% w/w Hydrogen peroxide stock solution (RICCA Chemical, catalog number: 3821.7-32 )
   
3. Sodium thiosulfate solution (dissolved in ddH₂O) (Sigma-Aldrich, catalog number: S8503 )
   
4. P.aeruginosa strains in freezer stock
   
5. 25% Lennox broth (LB) medium (Becton Dickinson and Company, Difco^TM, catalog number: 240230 ) (see Recipes)
   
6. LB agar plates (see Recipes)

Equipment

1. 96-well plates
   
2. Standard petri plates
   
3. Spectrophotometer (cuvette) (Thermo Fisher Scientific, model: GENESYS 10S UV-Vis )
   
4. Spectrophotometer (96-well plate) (Bio-Rad Laboratories, model: 680 )
   
5. Cuvettes for OD₆₀₀ reading
   
6. Shaker-incubator at 37 °C, 250 rpm
   
7. Static incubator at 37 °C
   
8. Sterile glassware: 150 ml Erlenmeyer flask, capped or foiled
   
9. Sterile 15 mm glass test tubes and plastic caps
   
10. Sterile wire-loops (sterilized with 70% ethanol and flame)

Procedure