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Quantitative RT-PCR analyses of GAPdH, VZV ORF63 and ORF68 expression in VZV-infected monocytes, NK cells, NKT cells, B cells, CD4+ T cells, CD8+ T cells and HFLs.

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posted on 14.03.2019, 13:40 by Dallas Jones, Christina N. Como, Lichen Jing, Anna Blackmon, Charles Preston Neff, Owen Krueger, Andrew N. Bubak, Brent E. Palmer, David M. Koelle, Maria A. Nagel

Human PBMCs were infected with VZV as described in Fig 1, then uninfected- and VZV-infected monocytes, NK cells, NKT cells, B cells, CD4+ T cells and CD8+ T cells were sorted using flow cytometry. RNA was harvested and then analyzed by quantitative RT-PCR for ORF63 (grey bars), ORF68 (red bars) and GAPdH (green bars) transcripts. In addition, VZV-infected HFLs that were >90% VZV gE+ based upon flow cytometry analyses were harvested as a positive control for viral transcript expression. Results representative of sorted PBMCs from 3 different healthy individuals and from 3 independent HFL infections, with bar graphs representing average 1/Ct ± SD. Dotted line represents threshold for Ct values (1/36). All immune cell populations were >93% pure based upon flow cytometry sorting.

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