Pe conjugated anti human cd163 antibody

Cell components in peritoneal washes were re-suspended in RPMI-1640 supplemented with 10% FBS. Then, the total number of viable cells was counted, and the cells were infected with TelomeScan at 1 MOI for 24 hours at 37ºC to visualize cancer cells as GFP-positive cells.^24–26 The cells were further stained with Alexa Fluor 647-conjugated anti-human CD45 antibody (BioLegend, 304056, RRID: AB_2564155), phycoerythrin (PE)-conjugated anti-human CD14 antibody (BioLegend, 301805, RRID:AB_31418), and PE-conjugated anti-human CD163 antibody (BioLegend, # 333606, RRID:AB_1134002) as markers of leukocytes, macrophages, and TAMs, respectively. Nuclei were stained with 4ʹ, 6-diamidino-2-phenylindole (DAPI). Immunofluorescent staining was observed under an inverted fluorescence microscope (IX71; Olympus).

The samples were washed with PBS containing 2% FBS and centrifuged at 300 × g, 4°C for 5 minutes. Dead cells were stained with APC-Cy7-conjugated Zombie NIR (BioLegend, #423105), and the cells were subsequently stained with the following surface markers for 30 minutes at 4°C: Alexa Fluor 647-conjugated anti-human CD45 antibody (BioLegend), FITC-conjugated anti-human CD14 antibody (BioLegend, #301803, RRID:AB_314185), and PE-conjugated anti-human CD163 antibody (BioLegend). As the controls, IgG-isotype control antibodies (BioLegend, #400112) were used. Cells were analyzed by flow cytometry (FACS Array, Becton Dickinson), and data were reanalyzed with Flow Jo software (Tree Star).