2.4. Flow cytometry
For cell-surface staining, the lentivirus-infected T cells were
harvested and washed with
phosphate buffered saline (PBS)
followed by incubation with 1ug/ml recombinant protein TRIL-R1-Fc (R&D
Systems, Minneapolis, MN) at room temperature for 20 min. After being
washed, the cells were incubation with 1ug/ml APC conjugated anti-human
IgG secondary antibodies and other cocktail antibodies (Biolegend) at
room temperature for 15 min in the dark. After being washed with PBS,
the cells were analyzed with a BD celesta (BD Biosciences) software by
flow cytometry. The following monoclonal antibodies were used with the
indicated subtypes: APC-lableled human IgG-Fc (clone HP6017),
Percy5.5-lableled anti-CD4 (clone RPA-T4), Alex700-lableled anti-CD8
(clone HIT8a), BV421-lableled anti-CCR7 (clone G043H7), PE-lableled
anti-CD45RO (clone UCHL1), AF488-lableled anti PD-1(clone EH12.2H7),
BV605-lableled anti-LAG-3 (clone 11C3C65), BV785-lableled anti-TIM-3
(clone F38-2E2). All sample data analyzed was done on≥10,000 events
using the FlowJo V 10 data analysis software.