Materials and Reagents

1. 100 ml peripheral blood obtained via subcutaneous venepuncture
   
2. Heparin - to prevent clotting of blood
   
3. Histopaque-1077 (Sigma-Aldrich, catalog number: 10771 ) or any equivalent ficoll solution (density 1.077 g/ml)
   
4. Fetal calf serum (FCS)
   
5. PBS (chilled)
   
6. DC isolation kit (Miltenyi Biotec, catalog number: 130-094-487 )
   
7. Antibodies (BD biosciences unless stated otherwise)
   
   1. Anti-TCRa/b-FITC (catalog number: 347773 ) - isotype with mouse IgG₁-FITC
      
   2. Anti-CD19-FITC (catalog number: 553785 ) - isotype with IgG_2a-FITC
      
   3. Anti-CD14-FITC (catalog number: 561712 ) - isotype with IgG_2a-FITC
      
   4. Anti-HLA-DR-PE (catalog number: 556644 ) - isotype with IgG_2a-PE
8. MACS buffer (see Recipes)
   

Equipment

1. Magnetic-activated cell sorting (MACS) magnet
   
2. LS columns (Miltenyi Biotec, catalog number: 130-042-401 )
   
3. Centrifuge (50 ml tube capacity)
   
4. 50 ml polypropylene conical tubes (Falcon^TM)
   
5. 14 ml snap-cap polypropylene tubes
   

Procedure