2. Materials and methods
2.1. Virus and cells
The cytopathogenic HM-175 strain of HAV (ATCC VR-1402) was propagated and assayed in confluent FRhK-4 cells (courtesy of Prof. Albert Bosch, University of Barcelona). Semi-purified stocks were obtained from infected FRhK-4 cells by three freeze–thaw cycles after 8–11 days post-infection. Cell debris was pelleted at 660 × g for 30 min. Infectious viruses were quantified by determining the 50% tissue culture infectious dose (TCID50) with eight wells per dilution and 20 μl of inoculum per well (Pintó et al., 1994).