Surface and interior images of fixed NIH 3T3 cells treated with fluorescently labeled transferrin and lysotracker were taken at 100X magnification at three separate time points (0 min. incubation, 30 min. incubation, 90 min. incubation) in order to capture endosome migration over time (Figure 1). At t=0 min., it is clear that there is an abundance of transferrin molecules at the surface of the cell (Figure 1A), resulting in a relative high average total fluorescence intensity compared to the interior of the cell (2.5, 2.1 respectively) (Figure 2). At t=30 min. the average total fluorescence intensity of the surface and interior of the cell is relatively similar (3.3, 3.2, respectively) (Figure 2), At t= 90 min. there is a relatively higher average total fluorescence intensity at the interior of the cell compared to the surface of the cell (4.1, 3.3, respectively) (Figure 2). Additionally, it can be visually noted that there is a significantly larger presence of transferrin molecules in the interior image than the surface image (Figure 1H, 1G, respectively). Lysozymes are unable to be visualized due to overpowering green stain in composite images. 3D projections of the cells were unable to be created due to immense technical difficulties.
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