Materials and Reagents

1. Staphylococcus aureus subsp. aureus USA300
   
2. Dry ice
   
3. Dimethyl sulfoxide (DMSO) (Thermo Fisher Scientific, catalog number: D128-1 )
   
4. Polyclonal anti-Hla antibody (Abcam, catalog number: ab15948 )
   
5. Anti-alpha hemolysin (Hla) antibody conjugated to horseradish peroxidase (Abcam, catalog number: ab15949 )
   
6. 3,3’,5,5’ tetramethylbenzidine (TMB) (Sigma-Aldrich, catalog number: T4444 )
   
7. Stop reagent (Sigma-Aldrich, catalog number: S5689 )
   
8. Luria broth (LB) agar plates
   
9. Phosphate buffered saline (PBS) (pH 7.2) (Sigma-Aldrich, catalog number: P5119 )
   
10. 10 mg/ml Bovine serum albumin (BSA) in PBS (Sigma-Aldrich, catalog number: P3688 ) (see Recipes)
    
11. 0.05% Tween 20 in PBS (Sigma-Aldrich, catalog number: P3563 ) (see Recipes)
    
12. Trypticase Soy Broth (TSB) (see Recipes)
    

Equipment

1. -80 °C freezer
   
2. 14 ml polypropylene culture tube (BD, catalog number: 352059 )
   
3. Cotton ball or spill resistant caps
   
4. Microtiter 96 well EIA/RIA plates (Corning, Costar^TM, catalog number: 9017 )
   
5. Microplate reader (Molecular Devices, model: SPECTRA Max M2), or any other model capable of measuring a 96 well microtiter plate at OD 650 nm
   Note: If a different model of microplate reader is used, ensure appropriate model of microtiter 96-well plate is used as the model above may not work.
   
6. 37 °C shaker
   
7. Nitrile gloves
   
8. Multichannel pipetman (Eppendorf)
   
9. 0.22 µm syringe filter (PVDF) (Thermo Fisher Scientific, Fisherbrand^TM, catalog number: 09-720-3 )
   
10. 3 ml luer lock syringe (BD, catalog number: 309657 )
    
11. 1.5 inch needle (BD, catalog number: 305187 )
    
12. 2 ml cryotubes
    
13. Plastic wrap
    
14. Disposable inoculation loops (10 µl, PS, sterile, yellow) (LPS, catalog number: M122002 )
    

Software

1. Microsoft Excel or other data processing program

Procedure