Fitc conjugated igg1

The MSCs were harvested and isolated from a human umbilical cord’s Wharton’s jelly tissue [41 (link)]. The cells were cultured in a high glucose Dulbecco’s Modified Engle’s Medium (DMEM) (Gibco, Thermo Fisher, Waltham, MA, USA), which was supplemented with 10% FBS, 1% (v/v) antibiotics (100 U/mL P/S), and 1% sodium pyruvate. After reaching confluency under the appropriate conditions (37 °C, 5% CO₂), the cells were collected with 0.05% trypsin-EDTA for the follow-up experiments.
To characterize the phenotypes of the MSCs, the cells were harvested and detached with 2 mM EDTA in phosphate-buffered saline (PBS), then washed by PBS with 2% bovine serum albumin (BSA) and 0.1% sodium azide (Sigma-Aldrich, Burlington, MA, USA). Further, the cells were incubated with antibodies conjugated with fluorescein isothiocyanate (FITC) or PerCP-Cy5-5-A. The indicated markers were CD34-FITC, CD45-FITC, CD44-PE, and CD90-PerCP-Cy5-5-A (BD Pharmingen, San Diego, CA, USA). FITC-conjugated IgG1 and PerCP-Cy5-5-A-conjugated IgG1 (BD Pharmingen) were demonstrated as isotype controls. The specific surface antigens of MSCs were detected by a flow cytometry [42 (link)]. Then, the cells were analyzed by FACS software (Becton Dickinson LSR II, Canton, MA, USA). The eighth passage of the MSCs was used in the current study.

The MSCs were acquired from the Wharton’s jelly tissue of a human umbilical cord which were kindly gifted by Prof. Woei-Cherng Shyu [33 (link)]. The cells were cultured in an H-DMEM medium (Invitrogen), supplemented with 10% FBS (1% (v/v) antibiotics 100 U/mL P/S and 1% sodium pyruvate) and removed with 0.05% trypsin–EDTA after reaching confluence (37 °C, 5% CO₂). Vascular Endothelial Growth Factor (VEGF, 50 ng/mL, Prospec-Tany TechnoGene Ltd., Rehovot, Israel) was used to investigate endothelial differentiation capacity.
The specific surface antigen of the MSCs was characterized by flow cytometry [34 (link)]. The MSCs were incubated with antibodies conjugated with Fluorescein Isothiocyanate (FITC) and Phycoerythrin (PE) using the markers: CD14-FITC, CD45-FITC, CD44-PE, and CD105-PE (BD Pharmingen, San Diego, CA, USA). Isotype controls were demonstrated by PE-conjugated IgG1 and FITC-conjugated IgG1 (BD Pharmingen). FACS software (Becton Dickinson LSR II, Canton, MA, USA) was used to analyze the phenotype of the MSCs. Cells at 8th passages were used in this study.

The MSCs used in the current research were kindly supported by Prof. Woei-Cherng Shyu, which were acquired from the Wharton’s jelly tissue of a human umbilical cord [39 (link)]. The cells were cultured in high glucose Dulbecco’s modified Eagle’s medium (H-DMEM, Invitrogen) supplemented with 10% FBS, 1% (v/v) antibiotics (100 U/mL penicillin/streptomycin) and 1% sodium pyruvate.
The specific surface antigens of the MSCs were characterized through flow cytometry [40 (link)]. The MSCs were harvested and detached with 2mM EDTA in phosphate-buffered saline (PBS) and washed with PBS containing 2% bovine serum albumin (BSA) and 0.1% sodium azide (Sigma-Aldrich, Burlington, MA, USA). Then, the MSCs were incubated with antibodies conjugated with fluorescein isothiocyanate (FITC), phycoerythrin (PE) or PerCP-Cy5.5 against the indicated markers: CD14-FITC, CD34-FITC, CD45-FITC, CD44-PE, CD90-PerCP-Cy5.5, and CD105-FITC (BD Pharmingen, San Diego, CA, USA). Further, PE-conjugated IgG1 and FITC-conjugated IgG1 (BD Pharmingen) were applied as isotype controls. Ultimately, the MSCs were analyzed by FACS software (Becton Dickinson LSR II, Canton, MA, USA). The cells at the 8^th passage were used in the current research.