CCR7 expression on NK cells is because of membrane uptake from donor cells rather than expression of the endogenous CCR7 gene. (A) RT-PCR analysis of CCR7 expression in NK cells. The donor cells Clone9.CCR7 were positive for CCR7 transgene mRNA expression and served as negative control for endogenous CCR7 transcript, and T cells served as positive control. CCR7⁺ NK and CCR7⁻ NK cells both lacked the expression of endogenous CCR7 transcript over the entire 8-hour period of coculture. GAPDH was used as control for quality of cDNA synthesis. (B) Fluorescence microscopic analysis of CCR7 uptake. After coculture of NK and Clone9.CCR7 cells for 15 minutes, the cells were stained with an anti-CCR7 FITC Ab (Clone9.CCR7 cells, in green) and with anti-CD16 and anti-CD56 PE Abs (NK cells, in red), spun onto glass slides with the use of cytospin and mounted slides using Vectashield mounting media, and visualized CCR7 uptake by NK cells with the use of a fluorescence microscope (Nikon Eclipse TE2000-U) equipped with Coolsnap HQ camera/Roper Scientific photometries and Metamorph image acquisition software.