Fig. 4.

SHIP-1 and SLP-76 downregulation in obinutuzumab-experienced NK cells. Primary cultured NK cells were stimulated (2:1) for 18 h with biotinylated rituximab (RTX-exp)-, obinutuzumab WT (GA101 WT-exp)-, obinutuzumab (GA101-exp)-opsonized or not opsonized Raji (Ctrl-exp) and immunomagnetically purified by negative selection. a Relative SHIP-1 or SOCS-1 mRNA levels were measured by RT-qPCR. The fold change expression relative to Ctrl-exp population (set to 1) after normalizing with the GAPDH endogenous control is reported in the bar graph. Data (mean ± SEM) from n = 15 donors of three independent experiments are shown. ****p < 0.0001, ***p = 0.0001. b Western blot analysis of equal amounts of total lysates. The same membrane was immunoblotted as indicated. One representative experiment is shown (left panels). The relative values of SHIP-1 or SLP-76 were obtained by normalizing to the levels of Akt and are expressed as fold change respect to control population (set to 1). Data (mean ± SEM, n = 8) from three independent experiments are depicted in bar graphs, **p = 0.0078, *p = 0.0156 (right panels)