Anti cd105 pe

The 3rd–5th passages of MSCs were harvested by trypsinization and incubated with 10 μL of the following mouse anti-human antibodies: anti-CD34-FITC (BD Pharmingen, USA), anti-CD45-PE (BD Pharmingen, USA), anti-CD90-FITC (AbD Serotec, USA), anti-CD73-PE (BD Pharmingen, USA), and anti-CD105-PE (Miltenyi Biotec, Germany) for 30 minutes at 4°C in the dark. After incubation, cells were washed twice with PBS and fixed with 300 μL 1% (v/v) paraformaldehyde. The expression profiles of cell surface markers were then determined by FACSCalibur flow cytometry (Becton Dickinson, USA) using CellQuest software. Cells labeled with FITC-conjugated mouse IgG1 (eBioscience, USA) and PE-conjugated mouse IgG1 (eBioscience, USA) served as negative controls.

Transduced and parental wild type (wt) cells were stained with anti-CD45/VioBlue, anti-CD33/PE, anti-CD90/VioBlue, anti-CD105/PE and anti-CD73/APC (all Miltenyi Biotec, Bergisch-Gladbach, Germany) monoclonal antibodies (mAbs) to analyze the hMSC marker profile. To monitor for transgenic 4-1BBL expression, SCP-1 cells were stained with an anti-CD137L/PE (BD Bioscience, Heidelberg, Germany) mAb. Samples were analyzed using a MACSQuant Analyzer and the MACSQuantify software (both Miltenyi Biotec).

The cells were harvested and immediately processed for flow cytometric analysis as described in our previous study [23 (link)]. Briefly, hMSCs (passages 3^rd-5^th) were washed twice with PBS. 4 × 10⁵ hMSCs were then resuspended in 50 μl PBS, incubated with 10 μl fluorochrome-labeled mouse anti-human monoclonal antibodies: anti-CD45-FITC (BD Pharmingen, USA), anti-CD34-PE (Biolegend, USA), anti-CD90-FITC (AbD Serotec, USA), anti-CD73-PE (BD Pharmingen, USA), and anti-CD105-PE (Miltenyi Biotec, Germany) for 30 minutes at 4°C in the dark. After being incubated with the antibodies, cell pellets were washed twice with PBS and fixed with 1% (w/v) paraformaldehyde in PBS. Flow cytometry was performed by FACSCalibur™ Flow cytometer using CellQuest™ software (Becton Dickinson, USA).