FIG. 4.

GD1a must be added before, and not after, incubation of cells with Py to stimulate infection. (A) A1-1 cells were treated with 80 μM GD1a at the indicated time points with respect to addition of cells with Py. At 48 h after incubation of cells with Py, cells were subjected to immunofluorescence with an antibody against the large T antigen. Data represent means ± standard deviations of the results of at least 2 independent experiments. A total of 3 of 4,004 control cells expressed T antigen. (B) A1-1 cells expressing CFP-Rab7 were incubated first with BODIPY-GD1a at 4°C for 15 min and then with labeled Py at 37°C for 3 h. GD1a (80 μM) was added pre- or postinfection. The extent of colocalization between labeled Py and BODIPY-GD1a in the CFP-Rab7 late endosomes was quantified from at least 3 cells. Data represent means ± standard deviations. (C) Data are as described for panel A, except NIH 3T3 cells were used. A total of 78 of 1,006 control cells expressed T antigen.