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. 2007 May 2;2(5):e415. doi: 10.1371/journal.pone.0000415

Figure 3. Gag/Δpp65 expressed in yeast assembles into VLP chimeras strongly activating CD8+ memory T cells in human whole blood.

Figure 3

(A) Western blot of Gag/Δpp65 (lane 1) and Gag (lane 2) expressed in yeast and partially purified as sucrose cushion pellet after ultracentrifugation. Aliquots (20 µl each) of the indicated VLP preparation were subjected to SDS-PAGE followed by Coomassie-Blue staining and western analysis probed with anti-Gag and/or anti-pp65 [M, full range rainbow marker, Amersham]. (B) Frequencies of antigen-specific CD4 and (C) CD8 T cell activation after stimulation by sucrose cushion-purified yeast Gag and Gag/Δpp65 particles. Activated T cells were identified as CD69/IFN-γ double-positive lymphocytes by flow cytometry. Antigen samples were added to whole blood from HCMV seropositive donor 1. A VLP-free sample containing PBSE buffer was included as negative control (NC), a lysate from HCMV-infected fibroblasts served as positive control (PC). The threshold of significant T cell responses (0.05% of counted lymphocytes [15]) is indicated as dashed line.