FIG. 5.

Results of three-color fluorescence assay to monitor fusion process. FAST-DiI (red) and calcein-AM (green) were used to label MAGI-CCR5 target cells, while EBFP (blue) marked Env-expressing cells. (A) Distribution patterns of three different fluorophores were examined individually, and then colocalization was analyzed in the merge field. The HTC-1 mutant provides a negative control for fusion, the TM186 mutant forms smaller and fewer syncytia than the wild type, and the wild-type Env protein (WT) mediates the formation of big syncytia. (B) Low-magnification view of merged fields for HTC-1, TM186, TM188, and wild-type Env.