Example 5
For FACS analysis of NKG2D ligand expression, tumor cells were stained with mouse NKG2D-Ig fusion protein (R&D systems) according to manufacturer's instruction. Cell-surface phenotyping of transduced primary T cells was determined by direct staining with APC-anti-CD3e (clone 145-2C11; Pharmingen, San Diego, CA), PE-anti-NKG2D (clone 16-10A1; eBioscience, San Diego, CA) and FITC-anti-CD4 (Clone RM4-5; Caltag, Burlingame, CA) monoclonal antibodies. Cell fluorescence was monitored using a FACSCALIBER™ cytometer. Sorting of NKG2D ligand-expressing cells was performed on a FACSTARM™ cell sorter (Becton Dickinson, San Jose, CA).