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. 2019 Jan 9;116(7):2577–2582. doi: 10.1073/pnas.1812632116

Fig. 5.

Fig. 5.

Characterization of IAV-QD(625+705) and visualization of IAV vRNP separation into individual parts at the AN region. (A) Schematic representation of IAV-QD(625+705). (B) TEM images. (Scale bar: 50 nm.) The white arrowheads indicate QD625, and the hollow arrowheads indicate QD705. (C) Immunofluorescence assay of IAV-QD(625+705) on coverslips. (Scale bar: 2 µm.) (D) Image of an infected cell. The nucleus boundary, labeled by Nup153-Venus, of a host cell is highlighted by a dashed line. The arrow marks the separation site. (Scale bar: 10 µm.) (E) Colocalization of QD625 and QD705. (F) Trajectories of QD625 and QD705. (G) Snapshot of the real-time separation of the QD625-vRNP and QD705-vRNP from D. (Scale bar: 1 µm.) (H) Differential interference contrast (DIC) image of an infected cell. The nucleus boundary is highlighted, and the arrow marks the uncoating site (red star). (Scale bar: 10 µm.) (I) Snapshots of the separation of QD625-vRNP (red), QD705-vRNP (green), and QD525-viral envelope (cyan). After the shift of cyan from green and red, the green and red were separated from each other. (Scale bar: 2 µm.) (Insets) Unmerged panels for QD625-vRNP and QD705-vRNP. Arrowheads indicate the separation of three QD signals. (J) Diagram of the two regions of cytoplasm. (Scale bar: 10 µm.) (K) Analysis of IAV vRNP separation in the AN region with and without drug treatment. A total of 1,017 trajectories in drug-treated cells and 1,130 trajectories in untreated cells were analyzed. **P < 0.01. p.i., postinfection. Images were taken at 60× magnification objective lens under a confocal microscope.