Fig. 2.

Characterization of YK614 extracellular particles. Vero cells were infected for 22 h with YK614; then cell culture supernatants were harvested, and cell debris was removed by low-speed centrifugation at 1,000 × g for 15 min, followed by 10,000 × g for 5 min. These supernatants were incubated with polylysine- and laminin-coated glass coverslips for 16 h in a humidified incubator at 37°C. Virus particles that stuck to coverslips were fixed with 4% paraformaldehyde, permeabilized with 0.2% Triton X-100, and then imaged by deconvolution microscopy as described previously (22). (A) mRFP-gB fluorescence superimposed on VP26-Venus fluorescence. (B and C) VP26-Venus and mRFP-gB fluorescence, respectively. (D) mRFP-gB fluorescence enhanced by raising the minimum intensity that was assigned maximum fluorescence, so that the red puncta numbered 3 and 5 were more intense than those in panel C. (E to G) Histograms of the number of pixels on the y axis versus the relative pixel intensity on the x axis for panels B to D. Panel G differs from panel F in that any pixel to the right of the diagonal red line has been assigned maximum intensity. VP26-Venus puncta 1 to 5 are labeled in all the images.