FIGURE 3.

Incubation of HD5 and BKV inhibits viral binding. A, AF488-BKV (m.o.i.: 16) was incubated with H₂O(black line) or 50 μg/ml of the indicated defensins (dotted line) in MEM, MEM containing 5% FBS, or PBS at 4 °C for 1 h. Vero cells in suspension were then incubated with the BKV mixture at 4 °C for 1 h and viral binding was evaluated using flow cytometry. Filled histograms represent cells in the absence of AF488-BKV. B, a 1:100 dilution of purified BKV was incubated with H₂O or 50 μg/ml HD5 in the indicated media at 4 °C for 1 h. This preparation was 2-fold serially diluted across a U-bottom plate containing PBS. The HA plate was incubated with 0.5% erythrocytes in PBS at 4 °C for 16 h. A representation image of a HA plate is shown. C, Vero cells in suspension were treated with 50 μg/ml HD5 (dotted line) or an equal volume of H₂O(bold line) in the indicated solutions at 4 °C for 1 h. Cells were washed and incubated with AF488-BKV (m.o.i. 16), biotintylated-SNA (lectin specific for α(2,6)-linked sialic acid), or biotintylated-MALII (lectin specific for α(2,3)-linked sialic acid) at 4 °C for 1 h. For SNA and MALII samples, cells were washed and incubated with AF488-labeled streptavidin. Fluorescence intensity was measured using flow cytometry. Filled histograms represent unstained cells.