Anti ly6g apc cy7 1a8

Cells were blocked in 100 μL of 20% normal mouse serum (collected in house) and 1 μg/mL anti-FcγII/FcγIII (2.4G2, Walter and Eliza Hall Institute) on ice for 20 min. Cells were stained with anti-CD45-PE.Cy5.5 or anti-CD45-Alexa 700 (30-F11), anti-CD11c-FITC (M5/114.15.2; eBioscience, San Diego, CA, USA), anti-CD11b-BV421 (M1/70), anti-CD19-BV510 (6D5), anti-CD4-BV650 (RM4-5), anti-CD103-BV786 (2E7), anti-CD64-PE (X54-5/7.1), anti-MHCII-PE.Cy7 (M5/114.15.2) and anti-Ly6G-APC.Cy7 (1A8; all from BioLegend, San Diego, CA, USA unless indicated). Cells were washed twice, resuspended in 0.25 mg/mL propidium iodide (ImmunoChemistry Technologies, Bloomington, MN, USA) and acquired on a BD LSRFortessa X-20 flow cytometer (BD Biosciences, San Jose, CA, USA). Single cells were identified as a linear population on FSC-A vs FSC-H then propidium iodide positive dead cells excluded. Cell counts were normalised using AccuCount Fluorescent Particles (Spherotech, Lake Forest, IL, USA). Data were analysed using FCS Express (De Novo Software, Los Angeles, CA, USA).