Cd107 pe lamp 1 antibody

The mice were euthanized and the spleen was surgically removed and placed in PBS. The single cell solution was obtained by passing the spleen through a 70 µm strainer. Splenocytes were separated by gradient centrifugation with Ficoll-Paque™ PLUS (VWR, Radnor, PA, USA) and red blood cells were removed using red blood cells (RBC) lysis buffer (eBioscience, 00-4333-57). Splenocytes were cryopreserved in FBS with 10% Dimethyl sulfoxide (DMSO) until functional experiments. For activation, splenocytes were co-cultured with Ab1 cells in a ratio 10:1. Staining was performed using IFNγ-PE antibody (BioLegend, 505807) and CD107-PE (LAMP-1) antibody (BioLegend, 121611) together with protein transport inhibition using Golgi Stop (BD Pharm, 554715). T cell proliferation was determined based on the dilution of CellTrace™ carboxyfluorescein succinimidyl ester (CFSE) dye (Thermo Fisher Scientific Corp.) and cytotoxic activity was measured using PrestoBlue™ Cell Viability Reagent (Thermo Fisher Scientific Corp.).

Balb/CByJ mice were euthanized and spleens were surgically removed and placed in PBS. Single-cell solution was obtained by passing an organ through 70-µm strainer. Splenocytes were separated by gradient centrifugation with Ficoll-Paque™ PLUS (VWR). Red blood cells were removed using RBC lysis buffer (eBioscience, 00-4333-57). Splenocytes were cryopreserved in FBS with 10% DMSO until functional experiments. For activation, splenocytes were co-cultured with Ab1 cells at a ratio of 10:1.
Staining was performed using IFNγ-PE antibody (BioLegend, 505807) and CD107-PE (LAMP-1) antibody (BioLegend, 121611) together with protein transport inhibition with Golgi Stop (BD Pharm, 554715). T cell proliferation was determined based on dilution of CellTrace™ CFSE dye (Thermo Fisher). CD4+ activation was analyzed using CD25-PE antibody (BioLegend, 102007) and CD134 (OX40)-BV421 antibody (BioLegend, 119411).