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. 2011 Mar 28;6(3):e18091. doi: 10.1371/journal.pone.0018091

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Sengupta et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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(A) Increased adenoviral E1A gene copy number in Ad-RGD treated cells (black bar) in comparison to Ad-WT treated cells (white bar) at different time-points after virus transduction calculated by qPCR. 24-fold higher viral E1A gene copy number was observed in Ad-RGD treated cells versus Ad-WT transduced cells at 24 h time-point (*p<0.001). (B) Bar diagram of flow-cytometric analysis showing percentage of T cells expressing viral hexon antigen at different time-points after virus transduction. Ad-RGD transduced T cells are represented by black bars and Ad-WT treated cells by white bars. (*p<0.0001; ns not significant). (C) Viral progeny was measured from cell lysate (cell-associated; upper panel) and culture supernantant (cell-free; lower panel) of adenovirus infected T cells. 16- to 21-fold increase in viral progeny was measured in T cell lysates and 64- to 77-fold higher viral progeny was observed in cell-free fractions of Ad-RGD treated cells (black bars) when compared to cells treated with Ad-WT (white bars) at different time-points after virus transduction (p<0.001; ND not detected). (D) Viability of virus-transduced T cells was assessed by Annexin/7AAD exclusion method. Frequencies of Annexin/7AAD double negative cells observed in flow-cytometry were plotted in a bar diagram. 1.8-fold and 2.8-fold higher death of Ad-RGD transduced T cells (black bars) was observed at 48 and 72 h after virus transduction respectively, when compared to Ad-WT transduced T cells (white bars). Mock (patterned bars) and Ad-WT showed similar viability through the entire time-course observation (*p<0.05; **p<0.001; ns not significant). Error bars represent mean ± SD.