Mouse igg2a fitc

Phenotypic analysis was carried out by FACS (fluorescence-activated cell system). Specifically, pCPC were immunostained by using specific monoclonal antibodies c-kit porcine-PE (DakoCytomation, Carpinteria, CA, USA), CD34-PE, CD45-FITC, CD90-FITC, and CD166-PE (BD Pharmingen, San Diego, CA, USA). Mouse IgG1-FITC (Caltag Medsystems, Buckingham, UK), mouse IgG2a-FITC, and rat IgG1-FITC (Abcam, Cambridge, UK) were used as isotype controls (Table S1). Quantitative analyses were performed using a flow cytometer FACS Scan (BD Biosciences, Franklin Lakes, NC, USA). Additional antibodies used for comparative expression analysis between pCPC and hCPC are also listed in Table S1.
Stably transduced pCPC populations with different lentiviral vectors were sorted by eGFP or mCherry fluorescence also with the FACS technique (flow cytometer FACS Aria; BD Biosciences).

Culture-expanded cells were washed with PBS-containing 0.3% (w/v) bovine serum albumin (BSA) and the concentration was adjusted to 1 × 10⁶ cells/100 μl. PDMSCs at passage 3, AL cells and DePDMSCs at passage 3 were examined for mesenchymal and hematopoietic marker expression of surface antigens by incubating with the antibodies CD14-phycoerythrin (PE), CD29-fluorescein isothiocyanate (FITC), CD44-FITC, CD105-PE and HLA-DR-FITC (Abcam, Cambridge, UK). Antibodies including mouse IgG2a-FITC, mouse IgG2a-PE/Cy5.5, mouse IgG1-FITC and rat IgG2b-FITC (Abcam) were used as isotype controls. After being labeled with antibodies in the dark at room temperature for 30 min, cells were washed twice with PBS. Flow cytometry was conducted using a BD LSR II (Beckman Coulter, Los Angeles, CA, USA), and the data were analyzed using BD FACSDiva software.

CD1a, SWC3a, CD80/86, and MHC-II on the surface of MoDCs were determined using flow cytometry (Beckman-Coulter, California, USA). Mouse anti-porcine FITC-CD1a antibody and FITC-anti-Monocyte/Granulocyte antibody (Abcam, Cambridge, UK) were used to detect DCs. Mouse anti-porcine FITC-SLA-DR antibody (AbD Serotec, Kidlington, UK) and the R-PE-CD152 (CTLA-4)-muIg (Ancell, California, USA) were used for the detection of DC maturation. Isotype control antibodies mouse IgG1-FITC, mouse IgG2a-FITC (Abcam, Cambridge, UK), mouse IgG2b-FITC (AbD Serotec, Kidlington, UK), and mouse IgG1-PE (Ancell, California, USA) were used for background control.