Cos-1 cells were transfected with Japan/57 NA, fixed at 48 hours post-transfection and processed for thin section immunogold labeling and electron microscopy as indicated in the Materials and Methods. Tetherin was detected with primary antibodies and labeled with secondary antibodies conjugated to 18 nm gold beads (panels A, B, and B1). NA was detected with primary antibodies and labeled with secondary antibodies conjugated to 12 nm gold beads (panel C) and 18 nm gold beads (panel D), respectively. Panel A, tetherin labeled at and near the plasma membrane. Panel B and inset B1, Japan/57 VLPs detected at the plasma membrane lacking tetherin immunogold colocalization. Panels C and D, Japan/57 VLPs labeled with 12 nm or 18 nm gold beads adjacent to the plasma membrane, respectively. Small arrows in panel A indicate tetherin gold labeling at and near the plasma membrane. Large arrows in panel D indicate NA gold labeling on VLPs at the cell surface.