Fig. 2

Confocal microscopy demonstrating TNTs traversing the pores of a transwell polyester membrane filter. MSTO cells were primed in TNT medium (low-serum, hyperglycemic medium) for 7 days prior to plating. To reduce exosomal trafficking, culture medium was removed 24 h prior to the experiment; cells were washed with PBS, and serum-free medium was added. Cells were stained with DiI prior to placement for culture on the top and bottom filters of a modified Boyden chamber encompassing a polyester filter containing pores measuring 400 nm in diameter. a DiI-stained cells with TNTs are shown on the left-hand panel. Comparison of MSTO cells and 400 nm pores is shown in the right-hand panel, with demonstration of Transmitted Detection (TD) imaging. Cells are marked by yellow cross bars and a pore is marked by an arrow head. b XZ view of Z-stacked confocal images of MSTO cells and associated TNTs/TNT-like structures crossing the membrane filter (indicated by arrowheads)