Intracellular IFN-α staining was performed according to the intracellular staining protocol from BD Bioscience. Cells were treated with Brefeldin A (GolgiPlug, BD Bioscience, 555029) 6 h prior to harvesting and stained with an anti-human IFN-α antibody (Miltenyi Biotec, 130–092-602). RPE cells and monocyte-derived cells were distinguished by the expression of CD206 (anti-human CD206, biolegend, 321124) or CD163 (anti-human CD163, BD Bioscience, 556018) on monocyte-derived cells as well as CellTracker Violet BMQC (Invitrogen, C10094) staining of monocyte-derived cells prior to co-culturing. Data were acquired on a LSRII flow cytometer (BD Biosciences) and analyzed with FlowJo software (Tree Star).
Anti human cd206
Manufactured by BioLegend
Anti-human CD206 is a monoclonal antibody that recognizes the CD206 antigen, also known as the macrophage mannose receptor. CD206 is a cell surface glycoprotein expressed primarily on macrophages and dendritic cells. It functions as an endocytic receptor, mediating the internalization of glycoproteins containing terminal mannose, fucose, or N-acetylglucosamine residues.
Automatically generated - may contain errors
Lab products found in correlation
Celltracker violet bmqc, by Thermo Fisher Scientific (1 mentions)
Anti human cd163, by BD (1 mentions)
Lsrii flow cytometer, by BD (1 mentions)
A00 1 1102, by Beckman Coulter (1 mentions)
Mouse anti cd68, by BioLegend (1 mentions)
Anti human cd86, by BioLegend (1 mentions)
Anti human cd68, by BioLegend (1 mentions)
Anti mouse f4 80, by BioLegend (1 mentions)
2 protocols using anti human cd206
1
Intracellular IFN-α Detection in Monocytes
2
Multiparameter Flow Cytometry Analysis
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After centrifugation and washing with PBS, surface flow cytometry antibodies were added according to the manufacturer’s instructions [anti-mouse CD11b (Biolegend, 101236), anti-mouse F4/80 (Biolegend, 353283), anti-human CD86 (Biolegend, 327702)], and incubated at 4°C in the dark for 30 min, then centrifuged the cells at 1500 rpm for 5 min.
Intracellular staining was added with 100 μL fixative solution at 4°C for 10 min, centrifuged at 1500 rpm for 5 min, discarded supernatant, then added 100 μL cell lysis buffer, and incubated at 4°C for 10 min. After washing with PBS, antibodies (anti-mouse CD68 (Biolegend, 333815), anti-mouse CD206 (Miltenyi Biotec, 130-102-604), anti-mouse FLT3 (Biolegend, 135310), anti-human CD68 (Biolegend, 333815) and anti-human CD206 (Biolegend, 354997)) were added and incubated in the dark at 4°C for 30 min. The cells were centrifuged at 1500 rpm for 5 min to discard unbound antibodies, resuspended in 100 µL PBS and tested by using a flow cytometer (Beckman Coulter, A00-1-1102).
Intracellular staining was added with 100 μL fixative solution at 4°C for 10 min, centrifuged at 1500 rpm for 5 min, discarded supernatant, then added 100 μL cell lysis buffer, and incubated at 4°C for 10 min. After washing with PBS, antibodies (anti-mouse CD68 (Biolegend, 333815), anti-mouse CD206 (Miltenyi Biotec, 130-102-604), anti-mouse FLT3 (Biolegend, 135310), anti-human CD68 (Biolegend, 333815) and anti-human CD206 (Biolegend, 354997)) were added and incubated in the dark at 4°C for 30 min. The cells were centrifuged at 1500 rpm for 5 min to discard unbound antibodies, resuspended in 100 µL PBS and tested by using a flow cytometer (Beckman Coulter, A00-1-1102).
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