text: CD4+ T-cell isolation by MACS
  Anti-CD4 microbeads were used essentially as recommended by the manufacturer (Miltenyi) [19]. PBMC were resuspended in 80 mul of FBS staining buffer. Anti-CD4 microbeads (20 mul) were added and incubated for 15 min at 6-12degreesC. Saturating amounts of fluorochrome-conjugated antibodies were added for a further 10 min. Cells were diluted in 2.5 ml of FBS staining buffer, pelleted, resuspended in 500 mul and magnetically separated, usually on an AutoMACS magnet fitted with a MACS MS column. Flow-through and two 1 ml washes were collected as the negative fraction. Enriched cells were collected in two 0.5 ml aliquots from the column after removal from the magnet. Alternatively, cells stained with anti-CD4-phycoerythrin were washed, magnetically labeled with anti-phycoerythrin microbeads (20 mul added to 80 mul of cell suspension; 15 min, 6-12degreesC), and magnetically separated as described above. The purity of cells was assessed by flow cytometric analysis of stained cells on a FACS Vantage sorter. Most (more than 97%) of the isolated cells had the CD4 T cell marker. 
  