Cytofix cytoperm fixation solution

Cells were fixed with Cytofix/Cytoperm Fixation solution (BD Biosciences) for 20 min at 4°C, then washed with Perm Wash Buffer/phosphate-buffered saline (PBS, 1 × ; BD Biosciences) and permeabilized with Perm Wash Buffer/PBS +0.1% Triton X-100 for 30 min. Cells were blocked with 3% BSA in 1 × Perm Wash Buffer at room temp for 30 min. After blocking, cells were incubated in primary antibody (Supplementary Table S2) diluted in 1 × Perm Wash Buffer +0.1% Triton X-100 4°C for 45 min. Alexa Fluor^®-tagged secondary antibody was added after primary incubation for 1 h at room temperature. Samples were run on a Beckman Coulter CyAn-ADP flow cytometer, and subsequent datasets were analyzed using FlowJo software.

Cells were incubated with Cytofix/Cytoperm Fixation solution (BD Biosciences) for 20 min at 4°C, then washed with Perm Wash Buffer/PBS (1×, BD Biosciences) and incubated in primary antibody for 2 h at room temperature. Cells were then incubated in Alexa Fluor-tagged secondary antibody for 1 h at room temperature. Samples were run on a Beckman Coulter CyAN_ADP flow cytometer. Datasets were analysed using FlowJo.