Pe conjugated anti human il 2rα

Unmodified YT-1⁵³ and
IL-2Rα⁺ YT-1 human NK cells⁵⁴ were cultured in RPMI complete medium
(RPMI 1640 medium supplemented with 10% fetal bovine serum, 2 mM L-glutamine,
minimum non-essential amino acids, sodium pyruvate, 25 mM HEPES, and
penicillin-streptomycin [Gibco]). CTLL-2 cells purchased from ATCC were cultured
in RPMI complete with 10% T-STIM culture supplement with ConA (Corning). 24
hours prior to signaling studies, CTLL-2 cells were resuspended in RPMI lacking
T-STIM culture supplement for IL-2 starvation. Viability (>95%) of CTLL-2
cells was verified by trypan blue exclusion (counted in a hemocytometer)
immediately before performing the signaling assays. All cells were maintained at
37°C in a humidified atmosphere with 5% CO₂. The subpopulation
of YT-1 cells expressing IL-2Rα was purified via magnetic selection as
described previously^{39 (link)}.
Enrichment and persistence of IL-2Rα expression were monitored by
analysis of PE-conjugated anti-human IL-2Rα (Biolegend) antibody binding
on an Accuri C6 flow cytometer (BD Biosciences).