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. 2024 Nov 10;13(22):1862. doi: 10.3390/cells13221862

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© 2024 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

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Phenotypic characterization of monocyte-derived macrophages and comparative analysis of in vitro macrophage-mediated phagocytosis following CD47 (B6-H12) antibody treatment. (A) CD14-positive monocytes are isolated and cultured in a macrophage medium, followed by activation with recombinant human M-CSF for 6 d. Single cells are stained and analyzed using the BD FACSVerse System. (B,C) KHOS/NP cells are labeled with CellTrace™ CFSE dye (KHOS-CFSE) and incubated with 10 μg/mL anti-CD47 antibody or IgG ISO-type antibody for 30 min at 37 °C. KHOS/NP-CFSE cells are co-cultured with differentiated macrophages for 2 h. The percentage of macrophage-mediated phagocytosis (CD14-high, CellTrace-high, and CD11b-high) is measured by flow cytometry. Data are recorded using BD FACSVerse and analyzed using FlowJo software. The percentages are presented as the mean ± standard error of the mean, and statistical significance is determined by comparison with the IgG ISO-type antibody (** p < 0.01).