Fitc conjugated anti cd45 antibody

The expression of specific surface markers of each batch of cultured MSCs was examined using flow cytometry. After trypsinization, 5 × 10⁵ MSCs in 50 μl of phosphate buffered saline (PBS) were incubated with 5 μl of antibodies including the PE conjugated anti-CD34 antibody (BioLegend, USA), FITC conjugated anti-CD45 antibody (BioLegend, USA), PE conjugated anti-CD73 antibody (BioLegend, USA), FITC conjugated anti-CD90 antibody (BioLegend, USA) and FITC conjugated anti-CD105 antibody (BD Bioscience, USA), for 30 min at 4 °C in the dark. After washing with PBS, the cells were fixed with 1% formaldehyde for 15 min. For each marker, at least 2 × 10⁴ labeled cells were acquired using flow cytometry (FACSCalibur, Becton Dickinson, USA). The data were analyzed by the CellQuest software version 3.3 licensed to Thammasat University (https://timothyspringer.org/files/tas/files/cellquest-softwarereference.pdf).

E0771 cells were stained with PE-conjugated anti-mouse CD284 (Biolegend, 117605) and Alexa Fluor 488-conjugated anti-mouse CD282 (Biolegend, 121807). To analyze CD8+ T cell infiltration in tumor tissues, E0771 tumors from WT and Bgn KO mice were dissected and digested with collagenase II and DNase I. Single-cell suspensions were stained with FITC-conjugated anti-CD45 antibody (Biolegend, 103115) and APC-conjugated anti-CD8a antibody (Biolegend, 100711). CD45 + CD8+ T cells were analyzed using a FACS Aria II. Data was analyzed using FlowJo V10 software (Tree Star Inc.).