Fig. 3.

Optimisation of pre-seeding and infection time periods. Cells were seeded either 6 or 16 h prior to infection, which was then allowed to proceed for 24, 48 or 72 h before fixation. The optimal confluency was obtained when cells were seeded at −6 h and fixed at 48 h post-infection (A); this correlated to the lowest background count of red cells per well in the GND controls (B) and is therefore the optimal time frame. Data represents the mean + SEM of titres calculated from three independently infected titre plates. (C) Representative blended images (bright phase-red fluorescence) of Huh-7 monolayers seeded and infected with JFH-1 at the indicated time points. The optimal confluency and most easily identifiable fluorescent cells are visible in the 6 h pre-seeding, 48 h infection duration panel (central left).