Cells were Fc blocked in 1:50 TruStain FcX PLUS (BioLegend, catalog no. 156604) in 2% FBS/PBS for 10 min, followed by a stain of at least 30 min of 1:200 CD11b-FITC (Thermo Fisher Scientific, catalog no. 11-0112-82), 1:400 CD117-PE (Thermo Fisher Scientific, catalog no. 12-1171-82), 1:200 Sca-1-PE/Cy7 (BioLegend, catalog no. 122514), 1:1000 Ghost Dye Red 780 fixable viability dye (Cell Signaling Technology, catalog no. 18452), and/or 1:200 lineage-FITC (BioLegend catalog no. 133301). Cells were then washed twice with 2% FBS/PBS and run on the flow cytometer. For compensation, UltraComp eBeads compensation beads (Thermo Fisher Scientific, catalog no. 01-2222-42) were stained at the same concentration for the same amount of time as the cells to be analyzed, washed twice with 2% FBS/PBS, and run on the flow cytometer. For viability dye compensation, an aliquot of cells was stained only with 1:1000 Ghost Dye Red 780 fixable viability dye (Cell Signaling Technology, catalog no. 18452) for the same duration as the cells to be analyzed, washed twice with 2% FBS/PBS, and run on the flow cytometer.
Greenwood D.L., Ramsey H.E., Nguyen P.T., Patterson A.R., Voss K., Bader J.E., Sugiura A., Bacigalupa Z.A., Schaefer S., Ye X., Dahunsi D.O., Madden M.Z., Wellen K.E., Savona M.R., Ferrell P.B, & Rathmell J.C. (2022). Acly Deficiency Enhances Myelopoiesis through Acetyl Coenzyme A and Metabolic–Epigenetic Cross-Talk. ImmunoHorizons, 6(12), 837-850.
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