Anti cd44 pe clone im7

ADSCs and LDSCs at passages 2, 4, and 6 were analyzed by flow cytometry. The cells were placed in fluorescence-activated cell sorting (FACS) tubes (BD Biosciences; 2 × 10⁵ cells/tube), washed with FACS buffer (PBS containing 2% FBS), and then incubated with the following fluorescein (FITC)- or phycoerythrin (PE)-conjugated antibodies: anti-CD14-FITC (clone M5E2; BD Pharmingen), anti-CD29-PE (clone TS2/16; BioLegend), anti-CD34-PE (clone 1H6; R&D Systems), anti-CD44-PE (clone IM7; BioLegend), anti-CD45-FITC (clone YKIX716.13; eBioscience), and anti-CD90-PE (clone YKIX337.217; eBioscience) or their respective isotype controls [11 (link), 12 (link)]. The cells were washed twice with FACS buffer and resuspended in 500 μl FACS buffer. Fluorescence was evaluated by flow cytometry using a FACSCalibur instrument (BD Biosciences). Data were analyzed using WinMDI 2.9 analysis software.

For immunophenotyping T lymphocyte memory subsets, the TruStain FcX™ antibody (Clone 96, BioLegend) was added to block the non-specific binding of antibodies. Subsequently, splenocytes were incubated with Zombie RED for 10 min at room temperature. Finally, cells were stained with the following fluorophore-labeled anti-mouse monoclonal antibodies: anti-CD4 FITC (clone GK1.5, BioLegend), anti-CD8 PCy7 (clone QA17A07, BioLegend), anti-CD44 PE (clone IM7, BioLegend) and anti-CD62L PerCP (clone MEL-14, BioLegend) for 20 min at 4 °C in the dark. The T lymphocyte subsets were characterized by the percentage values obtained from the SLA-stimulated culture.