Cell expansion was determined using a Cell Counting Kit-8 [24 (link)] (CCK8; Beyotime, Shanghai, China). NK cells were treated with calcitriol for 48 h and the supernatant was collected for the Cytokine-releasing assay. The cytokine levels were examined using the LEGEND Plex Human Inflammation Panel (BioLegend, California, USA). First, the cytokine capture beads were incubated with the standards or samples and then further incubated with biotinylated detection antibodies. The biotinylated detection antibody-binding solution, streptavidin (SA)-PE, was subsequently added to provide the fluorescent signal [25 (link)]. These signals were then analyzed using the FACS assay.
Cell cycle was measured using a Cell Cycle Detection Kit (Beyotime, Shanghai, China). NK cells were treated with calcitriol for 72 h. These cells were fixed in 70% ice-cold ethanol and then dyed with ribonuclease A (RNase A) and propidium iodide (PI) (Beyotime, Shanghai, China) [26 (link)].
Cell cycle was measured using a Cell Cycle Detection Kit (Beyotime, Shanghai, China). NK cells were treated with calcitriol for 72 h. These cells were fixed in 70% ice-cold ethanol and then dyed with ribonuclease A (RNase A) and propidium iodide (PI) (Beyotime, Shanghai, China) [26 (link)].
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