Quantifying Cell Proliferation by Flow Cytometry

Cell proliferation was assayed by Ki-67 (Alexa Fluor® 488 conjugate) antibody (Cell Signaling cat#11882). According to the manufacturer’s instructions, the cells after 7 days treatment are detached and washed with PBS. The cells were then fixed with 4% formaldehyde followed by incubation for 15 min at room temperature. After washing with PBS, cells were permeabilized using 0.1% PBS-tween on ice for 15 min. According to datasheet 1 × 10⁶ cells were incubated with the antibody for 1 h at room temperature after washing with PBS. Before measurement the cells were again washed 2 times and resuspended in PBS. The measurements were carried out using BD Accuri C6 Plus Personal Flow Cytometer (BD biosciences) and data processing was performed by FlowJo BD Accuri C6 Plus software for windows [27 (link)].