Pe conjugated anti human tim 3 mab

Cells were stained with antibodies, including APC/Cyanine7-conjugated anti-human CD14 mAb (325620, BioLegend, San Diego, CA, USA), PE/Cyanine7-conjugated anti-human CD163 mAb (333613, BioLegend), Pacific Blue-conjugated anti-human CD86 mAb (305423, BioLegend), 7-AAD Viability Staining Solution (420404, BioLegend), PE-conjugated anti-human TIM-3 mAb (345005, BioLegend) and PE-conjugated isotype control mAb (400111, BioLegend). Unstained samples were used as the negative controls. The stained cells were measured using a BD FACS Canto II flow cytometer (BD Biosciences, San Jose, CA, USA) and the data were analyzed using FlowJo software (version 10.8.1, FlowJo, Ashland, OR, USA).

Cells (HMC-1 cells and PBMCs) were incubated with human AB serum (Lonza) at 4°C for 5 min, and then stained with PE-conjugated anti-human TIM-1 mAb (Clone Name 1D12, BioLegend, San Diego, CA, USA), PE-conjugated anti-human TIM-3 mAb (Clone Name F38-2E2, BioLegend), PE-conjugated anti-human TIM-4 mAb (Clone Name 9F4, BioLegend) and PE-conjugated mouse IgG1 (Clone Name MOPC-21, BioLegend) at 4°C for 30 min. The expression of TIMs on the cells was determined with a FACS Canto II using Diva Software (BD Biosciences, San Jose, CA, USA).