Extended Data Fig. 1: Incorporation of RANK into mOC-SEVs.
From: Coupling of bone resorption and formation by RANKL reverse signalling
a, RANK content over time (top) in osteoclasts (lines) and their secretion products that were fractionated by serial centrifugation (columns) (n = 3); BLQ, below the limit of quantification. Bottom, osteoclast maturation over time with images of TRAP-stained cells and TRAP activity (n = 4). b, Transmission electron micrograph of precipitates from culture supernatants of mOCs by ultracentrifugation at 100,000g. Scale bar, 100 nm. c, RANK content in SEVs collected from osteoclast culture supernatants over time using ExoQuick-TC SEV-precipitating reagent (n = 3). d, Co-precipitation analyses of mOC-SEVs. Top, RANK content in co-precipitates (n = 3). Bottom, co-precipitates analysed for SEV markers (CD9, CD63, and CD81) and an endoplasmic reticulum protein (calnexin) by immunoblotting. GST pull-down (GPD) assays were used to precipitate RANK, and immuno-precipitation (IP) was used to precipitate CD9, CD63, and CD81, respectively. e, Confocal laser scanning micrograph of osteoclasts labelled for RANK (green), CD9 (red), and nuclei (blue). Scale bar, 20 μm. In a, c, d, data are mean, with individual points representing biologically independent samples.
