Anti nkg2a percp

Approximately 1 × 10⁶ cervical cancer cells per condition were seeded onto Petri dishes. Cells were incubated for 35 min at 4°C in staining buffer (PBS, 0.5% BSA, and 0.01% sodium azide). The cells were stained with specific antibodies against the following surface markers: anti-NKp30-PE, anti-CD95-APC, anti-CD39-FITC, anti-CD-73-PE-Cy7, anti-KIR3DL1-APC, anti-NKp46-APC, anti-CD25-PE, anti-CD25-PErCP-Cy5.5, anti-CD44-FITC, and anti-CD24-PE (Becton Dickinson, USA); anti-NKG2A-PerCP, anti-CD158e1-APC (R&D, Minneapolis, MN); and anti-CD158b2-FITC (BioLegend). Cells were washed and fixed with 1% paraformaldehyde for 20 minutes. For permeabilisation, cells were treated with Cytofix/Cytoperm buffer for 20 minutes (Becton Dickinson, USA). For intracellular staining, anti-human CTLA-4-APC, anti-CD95-PE-Cy7 (Becton Dickinson, USA), and anti-NKG2A-PerCP (R&D, Minneapolis, MN) antibodies were used. Cell samples were incubated for 35 minutes at 4°C in the dark. Finally, cells were washed and resuspended in staining buffer. The phenotype of tumor cells was characterized by flow cytometry on a FACSAria II cytometer (Becton Dickinson, USA). Data were analysed using Summit 4.4 software.