Materials and Reagents

1. RPMI-1640 medium (Life Technologies, Gibco^®, catalog number: 11875-093 )
   
2. FBS (Hyclone, catalog number: SH30396-03 )
   
3. Penicillin-Streptomycin-Glutamine Solution (Life Technologies, Gibco^®, catalog number: 10378-016 )
   
4. 2-mercaptoethanol (Life Technologies, Invitrogen™, catalog number: 21985-023 )
   
5. Phorbol 12-myristate-13-acetate (PMA) (Sigma-Aldrich, catalog number: 79346 )
   
6. Ionomycin (Sigma-Aldrich, catalog number: 19657 )
   
7. 5-(and-6)-Carboxyfluorescein Diacetate, Succinimidyl Ester (5(6)-CFDA, SE) (CFSE) (Life Technologies, Molecular Probes^®, catalog number: C1157 )
   
8. 7-hydroxy-9H-(1,3-dichloro-9,9-dimethylacridin-2-one (DDAO) (Life Technologies, Invitrogen™, catalog number: C34553 )
   
9. 5-(and-6)-(((4-Chloromethyl)Benzoyl)Amino) Tetramethylrhodamine (CMTMR) (Life Technologies, Invitrogen™, catalog number: C2927 )
   
10. 16% Paraformaldehyde Solution (PFA) (16% Formaldehyde, EM Grade) (Electron Microscopy Sciences, catalog number: 15710 )
    
11. Vectashield Hardset with Dapi (VWR International, catalog number: 101098-050 )
    
12. Poly-L-Lysine Solution 0.1% (w/v) in H₂O (Sigma-Aldrich, catalog number: P8920 )
    
13. SL8 (Ovalbumin (257-264)) (AnaSpec, catalog number: 60193 )
    
14. PBS (Life Technologies, catalog number: 14190-44 )
    
15. Media (see Recipes)
    
16. 2% PFA (see Recipes)
    
17. Poly-L-Lysine-coated chamber (see Recipes)
    

Equipment

1. Centrifuge
   
2. Flow Cytometer
   
3. Epifluorescent microscope
   
4. 37 °C, 5% CO₂ Cell culture incubator
   
5. 24 well plate (Corning, Costar^®, catalog number: 3524 )
   
6. 40 μm Cell strainer (BD Biosciences, Falcon^®, catalog number: 352340 )
   
7. FACS tube, polystyrene (BD Biosciences, Falcon^®, catalog number: 352054 )
   
8. 8-well chamber slide (Thermo Fisher Scientific, Lab-Tek^TM, catalog number: 177445 )
   

Procedure