Differentiation of Human-Induced Pluripotent Stem Cells into Macrophages
Published: July 31, 2023
Abstract
Source: Lopez-Yrigoyen, M., et al. Production and Characterization of Human Macrophages from Pluripotent Stem Cells. J. Vis. Exp. (2020).
This video describes an in vitro method for generating macrophages from human induced pluripotent stem cells (iPSCs). Macrophage generation from iPSCs occurs in three stages: (i) aggregation of iPSCs in suspension to form three-dimensional embryoid bodies (EBs) that differentiate into mesodermal cells, (ii) formation of macrophage precursor cells from the differentiated EBs, and (iii) differentiation of the macrophage precursor cells into mature, functional macrophages.
Protocol
1. Human iPSC Differentiation to Macrophages NOTE: A schematic summary of the macrophage differentiation protocol is depicted in Figure 1. Preparing cell differentiation growth factors and other reagents Prepare human embryonic stem cell-serum-free media (hESC-SFM; see Table of Materials) by supplementing Dulbecco's modified Eagle medium-F12 (DMEM/F12) with hESC supplement, 1.8% w/v…
Representative Results
Figure 1: Graphic summary of iPSC differentiation to mature functional macrophages. Diagram drawn with Biorender. Figure 2: iPSC differentiation towards macrophages and iPSC-DM number and morphology. (A) Bright Field images obt…
Disclosures
The authors have nothing to disclose.
Materials
| CellMask Deep Red Plasma Membrane Stain | Thermofisher | C10046 | Cryopreservation media |
| Cryostor CS10 | Sigma | C2874 | |
| CTS CELLstart Substrate | Invitrogen | A1014201 | Stem cell substrate |
| Porcine Skin Gelatin | Sigma | G9136 | |
| Recombinant Human BMP4 Protein | R&D | 314-BP-010 | |
| Recombinant Human IL3 | Preprotech | 0200-03-10 | |
| Recombinant Human MCSF (carrier-free) 100ug | Biolegend | 574806 | |
| Recombinant Human VEGF Protein | R&D | 293-VE-010 | |
| SCF (C-Kit Ligand) Recombinant Human Protein | Thermofisher | PHC2111 | |
| StemPro hESC SFM | Thermofisher | A1000701 | |
| StemPro EZPassage Disposable Stem Cell Passaging Tool | Thermofisher | 23181010 | |
| Ultralow attachment plates: Cell culture multi-well plate, 6 well, cell star cell repellent surface | Greiner | 657970 | |
| 2-Mercaptoethanol (50 mM) | Invitrogen | 31350010 | |
| DPBS, calcium, magnesium (500ml) | Thermofisher | 14040091 | |
| GlutaMAX Supplement | Thermofisher | 35050061 |
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Cite This Article
Differentiation of Human-Induced Pluripotent Stem Cells into Macrophages. J. Vis. Exp. (Pending Publication), e21516, doi: (2023).