Pe conjugated mouse anti human cd146

Briefly, PDLSCs were trypsinized and washed with PBS. Then, the cells were stained with primary antibodies including phycoerythrin (PE)-conjugated mouse anti-human CD34 (BD Biosciences, San Jose, CA, USA, catalog number: 555822), PE-conjugated mouse anti-human CD73 (BD Biosciences, catalog number: 550257), PE-conjugated mouse anti-human CD146 (BD Biosciences, catalog number: 550315) and fluorescein isothiocyanate-conjugated mouse anti-human STRO-1 (BioLegend, San Diego, CA, USA, catalog number: 340106) according to the manufacturer's protocols. Flow cytometry was performed on a BD Influx flow cytometer (BD Biosciences) and analyzed using a BD FACS™ Software version 1.0 (BD Biosciences).

Phenotypic analysis of hNSSCs was performed as described in our previous study (Vishnubalaji et al., 2012b (link)). Briefly, cells were washed twice in ice-cold PBS supplemented with 0.5% BSA and re-suspended at 10⁶ cells per ml. Ten microliters of PE-conjugated mouse anti-human CD146, CD73, CD29 and HLA-DR, FITC-conjugated mouse anti-human CD34, CD90, CD45, CD13, CD184 and CD31, or APC-conjugated mouse anti-human CD105, CD14 and CD44 antibodies (all from BD Biosciences, except the anti-human CD105, which was purchased from R&D systems) was added to 100 μl of cell suspension (105 cells). Negative control staining was performed using a FITC, PE, or APC-conjugated mouse IgG1 isotype control antibodies, respectively. Cells were incubated for 30 min at 4 °C in the dark, then were washed with PBS to remove excess antibodies, and then were resuspended in 500 μL of PBS and were analyzed using BD FACS Calibour flow cytometer (BD Biosciences. Data were analyzed using Cell Quest Pro Software Version 3.3 software (BD Biosciences).