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. Author manuscript; available in PMC: 2012 Mar 15.
Published in final edited form as: Cancer Res. 2011 Jan 31;71(6):2066–2076. doi: 10.1158/0008-5472.CAN-10-3200

Fig 3. T cells respond to NKG2D ligand positive cells by producing IFN-γ and exerting cytotoxicity in the presence of scFv-NKG2D.

Fig 3

Bulk spleen cells were stimulated with ConA (1ug/ml) and IL-2 (25 U/ml) before co-culture with irradiated tumor cells for 24 h. Both suspension (A) (105) and adherent tumor cells (B) (2.5×104) were co-cultured with ConA-stimulated spleen cells (105) in 96 well plates. Fusion protein scFv-huNKG2D (in which mouse NKG2D was replaced with the extracellular domain of human NKG2D) is used as negative controls. The final concentrations of scFv-NKG2D are 50 ng/ml. IFN-γ amounts in the supernatants were analyzed with ELISA. Results are shown in mean + SD. These data show that the expression of NKG2D ligands on tumor cells is required for induction of IFN-γ production. *: P<0.01 (scFv-mNKG2D vs ctrl). (C) ConA-stimulated T cells were co-cultured with NKG2D ligand-positive P815/Rae1 (mastocytoma), ID8 (ovarian cancer) and MC-38 (colon cancer) cells and –negative B16F10 (melanoma) at E:T ratios of 1:1 to 25:1 in the presence of scFv-mNKG2D (solid symbols) or control scFv-HuNKG2D (open symbols) of for 5h. The specific lysis was determined by Cr51-assay. The scFv-NKG2D proteins were added at concentrations of 10 and 100ng/ml. Results are shown in mean + SD of triplicates.