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Table 2 Detailed summary of the molecular markers and functional tests used to characterize microglia-like cells

From: Generating microglia from human pluripotent stem cells: novel in vitro models for the study of neurodegeneration

 Hematopoietic ProgenitorsMicroglia PrecursorsMicroglia-like CellsEfficiencyReproducibility
Protocol 1 – Muffat et al.Morphology: cystic structure
ICC: CD144 (CDH5), CD117 (KIT), CD41 (ITGA2B), CD235a (GYPA), PU.1 (SPI1)
Morphology: round, vacuolated cells, compact nucleus, filopodia, membrane ruffles
ICC: CD11b (ITGAM), IBA1 (AIF1), PU.1
Flow: CD11b, IBA1, CD45 (PTPRC)
Phagocytosis: latex bead assay
Proliferation: EdU assay
Morphology: multiple thin first-order branches terminated by membrane ruffles
ICC: TMEM119, P2Y12 (P2RY12), IBA1, CD45
Cytokines (CTRL, LPS + IFN-γ): Cytokine Array R&D
Transcriptional Profile: RNA-Seq
Yield: 0.5-4x
Purity: 97%
15 hiPSC-lines
3 hESC lines
Protocol 2 - Pandya et al.Morphology: floating cells
ICC: CD34, CD43 (SPN), CD45
not reportedFlow: CD11b, IBA1, CD45, CX3CR1, HLA-DR;
Negative: CD80, CD86, CD206, CD200R
Cytokines (CTRL, LPS): TNF-α, kit unspecified
Phagocytosis: pHrodo-E.coli
ROS: CellROX assay
Transcriptional Profile: Microarray
Yield: 0.8-3x
Purity: 9% (in mixed glial culture)
2 hiPSC lines
Protocol 3 –
Abud et al.
Morphology: endothelial cells
Flow: CD41 (ITGA2B), CD43, CD235a
ICC: PU.1, TREM2
Flow: CD11b (9%), CD45 (65%), CX3CR1 (22%); negative: CD117
Morphology: high nucleus to cytoplasm ratio
ICC: PU.1, CX3CR1, TREM2, P2Y12, TGFBR1, PROS1, MERTK, ITGB5
Flow: CD11bint, CD45low-int
Phagocytosis: pHrodo-E.coli, pHrodo-hS (CD11b-dependent, MERTK-independent)
Cytokines (CTRL, LPS, IL-1β, IFN-γ): V-PLEX cytokine 30-plex
Calcium Signaling: ADP-response (P2Y12-negated)
Transcriptional Profile: RNA-Seq
Transplantation: into MITRG mice
Yield: 30-40x
Purity: 97%
10 hiPSC lines
Protocol 4 - Douvaras et al.Morphology: adherent endothelial cells
Flow: CD309 (KDR), CD235a (up to 6%)
Morphology: floating cells
Flow: CD14, CD45, CX3CR1 (70%)
Morphology: ramified cells with motile processes
ICC: CD11c, IBA1, P2Y12, TMEM119
Flow: CD11b, CD11c, CX3CR1, P2Y12
Phagocytosis: latex beads
Cytokines (CTRL): Human XL Cytokine Array Kit
Calcium Signaling: ADP-response
Transcriptional Profile: RNA-Seq
Yield: 2x
Purity: 68% (CD14+)
14 hiPSC lines
2 hESC lines
Protocol 5 - Haenseler et al.Flow: CD34Morphology: large, vesicle-rich, floating cells
Flow: CD14, CD16, CD45, CD86, CD163, HLA-DR
Phagocytosis: pHrodo-zymosan
Morphology: ramified (secondary branches)
Flow: CD11b, CD11c, CD14, CD45, (CD33), (MERTK);
Negative: HLA-DR
Cytokines (CTRL, LPS/IFN-γ): Human XL Cytokine Array Kit (R&D), Luminex 100 Bio-Plex System (BioRad)
Yield: 10-43x
Purity: ~ 100% (CD14+)
4 hiPSC lines
Protocol 6 - Takata et al.not reportedMorphology: floating, round, vacuolated cells
Flow: CD11b, CD14, CD45, CD163, CX3CR1
CyTOF: CD11b, CD14, CD45, CD48, CD62L, CD64, CD115, CD163, CX3CR1, HLA-DR, MARCO, MERTK
Phagocytosis: latex beads, Aβ-TAMRA
Morphology: ramified cells
ICC: IBA1
FLOW: CX3CR1 (increased expression compared to d25)
Phagocytosis: latex beads, Aβ-TAMRA
Cytokines (CTRL, LPS): Human Magnetic Luminex Assay (R&D)
Transcriptional Profile: none
Yield: n.a.
Purity: 94% (CD14+/CD45+)
1 hiPSC line