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. 2014 Apr 19;456:268–278. doi: 10.1016/j.virol.2014.04.002

Fig. 4.

Fig. 4

Fig. 4

Confocal laser scanning microscopic examination of PEC entry. Confluent LLC-PK cells grown on Lab-Tek II CC2 chamber slides were infected either with mock (medium) or PEC (MOI 50) in the presence or absence of GCDCA (100 µM), fixed and stained at 1 h PI (A) or at 4 or 6 h PI (B) and observed under a confocal laser scanning microscope. (A) Top panels (a–d) show the mock-infected cells without GCDCA; middle panels (e to l) show the cells infected with PEC with or without GCDCA; bottom panels (m–p) show the cells infected with PEC in the presence of GCDCA and chloroquine. (B) Top panels (a–d) show the cells infected with PEC without GCDCA and observed at 4 h PI; bottom panels (e–h) show the cells infected with PEC without GCDCA and observed at 6 h PI. (C) Cells were infected with PEC (MOI 50) for 1 h, then treated with GCDCA for 5 min (a–d), 15 min (e–l), 30 min (i–l) or 60 min PI .(A–C) Cells were fixed and probed with rabbit polyclonal anti-Rab7 and swine polyclonal anti-PEC VLP primary antibodies and detected by PerCP-Cy5.5 labelled goat-anti-rabbit antibody (red) and FITC labelled goat-anti-swine antibody (green). Nuclei were stained with sytox orange (5 µM) (pseudo colored blue), and merged images for PEC and Rab7 were prepared. In the merged images, colocalization of PEC (green) and Rab7 (red) appears in white by using ImageJ.