Isolated and expanded MSCs were observed for plastic adherence and fibroblast-like spindle morphology. For flow cytometric characterization, 100 μl of MSC resuspended in culture medium at a concentration of 1–1,5 × 106 MSC/ml was incubated with 1,5 μl anti-CD73 [PE] (BioLegend cat. no. 344044), 1,5 μl anti-CD90 [APC-Cy7] (BioLegend cat. no. 328132), 5 μl anti-CD105+ [APC] (BioLegend cat. no. 323208), 0,5 μl anti-CD14 [FITC] (BioLegend cat. no. 301804), 0,5 μl anti-CD19 [FITC] (BioLegend cat. no. 302206) 0,5 μl anti-CD31 [FITC] (BioLegend cat. no. 303104), 1,5 μl anti-CD45 [FITC] (BioLegend cat. no. 368508), 0,5 μl anti-HLA-DR [FITC] (BioLegend cat. no. 307604) and 1 μl of 1 mg/mL 7-Aminoactinomycin D (7-AAD, AAT Bioquest, cat. no. ABD-17501) for 30 min at room temperature in dark. After incubation, 3 mL of PBS with 0.1% BSA was added to the cells, followed by centrifugation at 500 g for 5 min. The pelleted cells were resuspended in 400 μl of PBS with 0.1% BSA, followed by analysis on a Novocyte 3000 (Agilent). The acquisition rate was kept below 1.000 for all analyses. A fluorescence minus one-guided gating strategy was applied to identify positive and negative events. In all applied MSC batches, ≥90% of the viable cells expressed the positive markers CD73, CD90, and CD105, while ≤2% expressed the negative markers CD14, CD19, CD31, CD45, and HLA-DR.
Anti cd105 apc
Manufactured by BioLegend
Sourced in United States
Anti-CD105-APC is a cell surface marker antibody that binds to CD105, also known as endoglin, which is expressed on endothelial cells. This product is suitable for flow cytometry applications.
Automatically generated - may contain errors
Lab products found in correlation
Lsr 2 flow cytometer, by BD (3 mentions)
Anti cd34 pe, by BioLegend (3 mentions)
Anti cd19 fitc, by BioLegend (2 mentions)
Facsdiva software, by BD (2 mentions)
Isotype matched control antibody, by BD (2 mentions)
Anti cd45 apc, by BD (2 mentions)
Anti cd140b pe, by BioLegend (2 mentions)
Anti cd45 fitc, by BioLegend (1 mentions)
Anti cd14 fitc, by BioLegend (1 mentions)
Anti hla dr fitc, by BioLegend (1 mentions)
Novocyte 3000, by Agilent Technologies (1 mentions)
7 aminoactinomycin d 7 aad, by AAT Bioquest (1 mentions)
Anti cd73 pe, by BioLegend (1 mentions)
Anti cd31 fitc, by BioLegend (1 mentions)
Anti cd31 apc, by BD (1 mentions)
70 μm cell strainer, by Thermo Fisher Scientific (1 mentions)
Fitc isolectin b4, by Merck Group (1 mentions)
Anti cd31 pe, by BioLegend (1 mentions)
Murine igg, by Merck Group (1 mentions)
Legacy moflo mls high speed cell sorter, by Beckman Coulter (1 mentions)
10 protocols using anti cd105 apc
1
Immunophenotypic Characterization of MSCs
2
Characterization of C-MSC Surface Markers
3
Flow cytometry characterization of C-MSCs
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C-MSC were blocked with 5% rat serum and stained respectively with conjugated antibodies, including anti-CD105-APC (BioLegend, San Diego, CA, USA), anti-CD31-APC (BD Biosciences, San Jose, CA, USA), anti-CD45-APC (BD Biosciences), anti-CD140b-PE (BioLegend), or isotype-matched control antibody (BD Biosciences). Flow cytometry analysis of cultured C-MSC was performed with a BD LSRII flow cytometer and BD FACSDiva™ software (8.02, BD Biosciences, San Jose, CA, USA).
4
Isolation of Primary Lung Endothelial Cells
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Primary LMVECs were isolated as previously described (76 (link)). Briefly, lungs from female C57BL/6 mice were dissected, digested in a 0.5-mg/ml collagenase solution for 1 hour at 37°C, and filtered through a 70-μm cell strainer (Thermo Fisher Scientific). Suspended cells were washed, blocked with 10 μg/ml murine IgG (I5381, Sigma-Aldrich), and stained with isolectin-B4–FITC (L2895, Sigma-Aldrich), anti-CD31–PE (102408, BioLegend), and anti-CD105–APC (120414, BioLegend) antibodies, diluted to 1 μg/ml in PBS with 2.5% FBS. Isolectin-B4+CD31+CD105+ cells were sorted using a Beckman Coulter Legacy MoFlo MLS High Speed Cell Sorter and cultured.
5
Adipocyte Characterization Protocol
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IL-6, leptin and adiponectin ELISAs and human recombinant IL-6 were from R&D systems (Abingdon, UK). Anti-human antibodies (anti-CD31-FITC, anti-CD166-PerCP-efluor) were from eBioscience (Hatfield, UK), anti-CD105-APC, anti-CD45-Alexa fluor 700 and anti-CD11b-Brilliant Violet 421 were from Biolegend (Cambridge, UK) and anti-CD73-Brilliant Violet 605 was from BD Bioscience (Oxford, UK). Insulin ELISA was from Mercodia Diagnostics (Uppsala, Sweden). DAPI, LipidTOX Green Neutral Lipid, Inflammatory Cytokine Human Magnetic 5-Plex and Trizol were from Life Technologies (Warrington, UK). RT2 Profiler human adipogenesis PCR arrays and cDNA synthesis kits were from SABiosciences-Qiagen (Hilden, Germany). Human Oxidative Stress Magnetic Bead Panel was from Millipore (Watford, UK). Human white SC-pre-adipocytes from a lean individual (C-12730) were from PromoCell (Heidelberg, Germany). Other chemicals and reagents were from Sigma (Munich, Germany).
6
Isolation and Characterization of BMMSCs from OVX Mice
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BMMSCs were isolated from the femurs and tibias of sham-operated mice or OVX mice treated with PBS, TDNMCP-1, TDNFasL, or TDNs. Femurs and tibias were dissected out and cleaned of connective tissue. Cells were flushed out from long bones by a syringe with α-MEM supplemented with 20% FBS, 2 mM l -glutamine (Sigma-Aldrich, USA), and 1% penicillin/streptomycin (Invitrogen, USA). Single-cell suspensions were seeded in dishes and initially maintained in an atmosphere of 5% CO2 at 37 °C. The medium was changed every 3 days. After reaching 80% confluence, cells were passaged using 0.25% trypsin.
Cell surface markers of BMMSCs were detected by flow cytometry analysis. Cells were harvested and washed in PBS. Then, the single-cell suspension was incubated with mouse anti-CD105-APC (BioLegend, 120413), anti-CD11b-FITC (BioLegend, 101206), anti-CD45-PE (eBioscience, 12-0451), anti-CD29-APC (eBioscience, 17-0291-82) and anti-CD34-PE (BioLegend, 119307) antibodies. Finally, cells were washed twice in PBS, subjected to flow cytometry analysis with a flow cytometer (Cytomics FC 500; Beckman-Coulter) and analyzed with FlowJo_V10 software.
Cell surface markers of BMMSCs were detected by flow cytometry analysis. Cells were harvested and washed in PBS. Then, the single-cell suspension was incubated with mouse anti-CD105-APC (BioLegend, 120413), anti-CD11b-FITC (BioLegend, 101206), anti-CD45-PE (eBioscience, 12-0451), anti-CD29-APC (eBioscience, 17-0291-82) and anti-CD34-PE (BioLegend, 119307) antibodies. Finally, cells were washed twice in PBS, subjected to flow cytometry analysis with a flow cytometer (Cytomics FC 500; Beckman-Coulter) and analyzed with FlowJo_V10 software.
7
Characterizing iPSC-MSCs by Flow Cytometry
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Surface markers of iPSC-MSCs and NAMPT-iPSC-MSCs were evaluated by flow cytometry. Antibodies including anti-CD90-APC (BioLegend, B322310), anti-CD105-APC (BioLegend, 800507), anti-HLA-DR -APC/Cyanine7 (BioLegend, 307617), anti-CD73-APC (eBioscience, 17-0739-42), anti-CD34-PE (BioLegend, 4341649), anti-CD45-APC (BD, 560973) were used.
8
Quantifying Endoglin Expression in Glioma
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For the detection of endoglin in human glioma cell lines, we used anti-CD105-APC (BioLegend, San Diego, CA, USA) and an isotype-matched control antibody from eBioscience (San Diego, CA, USA). Specific fluorescence indexes (SFI) were calculated by dividing mean fluorescence obtained with the specific antibody by mean fluorescence obtained with isotype control antibody.
9
Umbilical Cord MSC Characterization
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UC−MSCs in MCB and WCB were recovered and seeded in T75 flasks at a cell density of 2 × 104 cells/cm2. When they reached 90–100% confluence, UC−MSCs were harvested for cell surface marker detection. For UC−MSC sheets, the sheet was digested using TrypLE into single-cell suspension for detection. UC−MSCs were aliquoted into 1 × 106 cells/tube in the staining buffer, consisting of phosphate buffer saline (PBS, Corning (Corning, NY, USA)), supplemented with 1% FBS (Gibco). Then, anti-CD73-FITC (BD (Becton, NJ, USA)), anti-CD90-FITC (BD), anti-CD105-APC (BioLegend (San Diego, CA, USA)), anti-CD11b-FITC (BD), anti-CD19-FITC (BioLegend), anti-CD34-PE (BioLegend), anti-CD45-FITC (BD), anti-HLA-DR-FITC (BD), anti-IgG-FITC (BD), anti-IgG-PE (BD), and anti-IgG-APC (BD) were added to the tubes separately. After staining for 30 min at room temperature in the dark, the cells were washed twice with PBS and resuspended in the staining buffer for flow cytometry analysis.
10
Cardiac Mesenchymal Stem Cell Immunophenotyping
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Cardiac mesenchymal stem cells were first blocked with 5% rat serum (Sigma) and stained, respectively, with conjugated antibodies, including anti-CD105-APC (BioLegend), anti-CD140b-PE (BioLegend), or isotype-matched control antibody (BD Biosciences). Flow cytometry analysis of cultured C-MSC was performed with a BD LSRII flow cytometer from Augusta University.
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