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. 2021 May 25;12(6):538. doi: 10.1038/s41419-021-03770-0

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — A Gating strategy to identify murine large (LPM) and small (SPM) peritoneal macrophages. Cells were gated as CD19⁻, Ly6G⁻ live cells. B Representative flow cytometric analysis for expression of AXL and MERTK on LPM and SPM. C Frequency of AXL or MERTK single and double positive LPM and SPM. Bar graphs show mean with SD and individual samples. D Experimental set-up to study efferocytosis in the murine peritoneum. Apoptotic cells (AC) were generated by treating murine thymocytes with dexamethasone. Afterwards, cells were labeled with pHrodo-Red and washed with EDTA/BSA and PBS. To set-up the model, initially different numbers of ACs were injected intraperitoneally. Peritoneal exudate cells (PECs) were analyzed at different time points by flow cytometry. Created with BioRender.com E Representative flow cytometric analysis of PECs for expression of F4/80 and pHrodo-Red signal. F Representative histograms for pHrodo-Red upon injection of different numbers of ACs and PECs isolated after 4 h (Gated on CD19⁻, F4/80⁺ live cells). Statistical analysis of pHrodo⁺ cells. Bar graphs show mean with SD and individual samples. G Representative histograms for pHrodo-Red upon injection of 2.5 × 10⁶ ACs and PECs were isolated at indicated time points (Gated on CD19⁻, Ly6G⁻, and F4/80⁺ live cells). Statistical analysis of pHrodo⁺ cells. Bar graphs show mean with SD and individual samples. H Representative histograms for pHrodo-Red upon injection of 2.5 × 10⁶ ACs with or without 50 nM GAS6. PECs were isolated at indicated time points (Gated on CD19⁻, Ly6G⁻, and F4/80⁺ live cells). Bar graphs show mean with SD and individual samples. Data were representative of two individual experiments (n = 3–5).