Figure 1.

Osteoclast preparation and cell sorting strategy. (A) Schematic representation of OCL preparation from mouse bone marrow progenitors and human peripheral blood mononuclear cells (PBMCs). (B) Gating strategy for FACS analysis and sorting of ≥3N cells using Hoechst 33342 after doublet discrimination. Nuclei numbers are indicated at the top of FACS graphs. (C,D) Representative cell sorting of 1–2N cells (orange) and ≥3N cells (blue) using flow cytometry for murine (C) and human (D) cells. After gating on single cells, 1–2N cells (P1, orange) and ≥3N (P2, blue) cells were selected for sorting. To validate the gating strategy and the sorting process, sorted cells were reanalyzed in a flow cytometer. Sorted cells with 1–2N and ≥3N revealed >99% positive events with corresponding nuclei number.