Figure 1

Cellular internalization of GapmeR delivered through direct uptake in human T-cells.

(A) Primary human T-cells were incubated with 100 nM, 250 nM or 500 nM non-targeting FAM-GapmeR for 6 h, 24 h, 48 h or 72 h and then GapmeR cellular uptake was analysed by flow cytometry. Results show dose- and time-dependent cellular internalization of GapmeR. (B) Both primary human T-cells and HuT78 cells were incubated separately with 500 nM FAM-GapmeR for up to 72 h and GapmeR cellular uptake was analysed by flow cytometry. Results (mean ± SEM) showed comparable rate of GapmeR internalization in HuT78 and primary T-cells. (C) Primary T-cells were incubated with 500 nM FAM-GapmeR (green) for 6 h, fixed and counter-stained with Rhodamine-Phalloidin (to visualize cells, red) and Hoechst (to visualized nuclei, blue). GapmeR cellular localization was analysed by confocal microscopy (63X oil objective lens). Images clearly show cytoplasmic and nuclear localization of GapmeR. (D) Time-dependent cellular internalization of FAM-GapmeR delivered through gymnosis (cell intensity and nuclear intensity) was quantified using High Content Analysis. Data (mean ± SEM) represent at least three independent experiments using T-cells purified from at least 3 different donors. Differences in nuclear/cell intensities between 24 h and 48 h treatments were non-significant (NS); *p < 0.05.