Fig. 5.

Results of Western blotting of culture supernatant, cell lysate, and purified recombinant measles viral particles. (A) Vero cells were infected with MVAIK, MVAIK/RSVF, and MVAIK/RSV/G and HEp-2 cells were infected with RSV subgroup A, Long strain, and were cultured in 1 ml in a 24-well plate. Just before the appearance of CPE, culture media was replaced with serum free medium (VP-SFM). 1 ml of culture medium was harvested and 100 μl of PBS was added in plate. Cells were freeze-thawed and cell lysate was clarified. As for the Western blotting, 1/30 of initial supernatants and 1/100 of cell lysate were subjected for experiments. They were stained with polyclonal antibodies against RSV. (B) Infectious particles were obtained through sucrose discontinuous gradient ultra-centrifugation. Fraction 1 was obtained at the top of the gradient of 30% sucrose, Fraction 2 between 30% and 45% sucrose, and Fraction 3 between 45% and 60% sucrose. Each fraction was analyzed by Western blotting, using RSV polyclonal antibodies and monoclonal antibodies against MV N protein.