Centrifugal Elutriation of Cell Cycle Phases

Centrifugal elutriation was performed using a Beckman JE-5.0 elutriation rotor as previously described (25 (link), 26 ) with slight modifications. Briefly, ALL cells (4–6 ×10⁸) were suspended in 25 ml of elutriation buffer (Hank’s buffered salt solution containing 1.6 g/L 2-naphthol-6,8-disulfonic acid dipotassium salt and 2% FBS), passed through a 25G needle twice, and introduced into the elutriation chamber at a flow rate of 25 ml/min with a rotor speed of 3000 rpm. Rotor speed was reduced to 2920 rpm to collect the W1 wash fraction and then to 2620 rpm (661g) to collect the F1 fraction containing cells in G1 phase. Successive reductions in rotor speed were performed to collect wash fractions, and cells in G2/M phases were collected in the F3 fraction at 1860 rpm (333g). Aliquots of the fractions were subjected to propidium iodide staining to verify DNA content. Cells were resuspended in fresh growth medium.