Figure 3.
Ultrafast SFMI of Cy3-labeled DOPE and TfR in the apical PM. (A) Schematic drawing showing that membrane molecules undergo hop diffusion in the PM, which is compartmentalized by the actin-based membrane-skeleton (MSK) meshes (fences; brown mesh) and rows of transmembrane-protein pickets anchored to and aligned along the actin fence (gray molecules). (B) A typical snapshot of single Cy3-DOPE molecules in the apical PM observed under our standard conditions, with an integration time of 0.1 ms. (C) A representative image sequence of a single Cy3-DOPE molecule diffusing in the apical PM, recorded every 0.1 ms (shown every 100 image frames = every 10 ms). The colors in the trajectory indicate the diffusion in different plausible compartments (order of appearance: purple, blue, green, orange, red, and then back to purple; the same color order is used throughout this report), detected by the TILD analysis software (Fig. S5). (D) Typical single-molecule trajectories of Cy3-DOPE and Cy3-Tf-TfR in intact and actin-depleted blebbed apical PMs, recorded at normal video rate and enhanced rates. The order of the compartments that the molecule visited (parenthesized integers) and the residency times there, as determined by the TILD analysis, are indicated (intact PMs).

Ultrafast SFMI of Cy3-labeled DOPE and TfR in the apical PM. (A) Schematic drawing showing that membrane molecules undergo hop diffusion in the PM, which is compartmentalized by the actin-based membrane-skeleton (MSK) meshes (fences; brown mesh) and rows of transmembrane-protein pickets anchored to and aligned along the actin fence (gray molecules). (B) A typical snapshot of single Cy3-DOPE molecules in the apical PM observed under our standard conditions, with an integration time of 0.1 ms. (C) A representative image sequence of a single Cy3-DOPE molecule diffusing in the apical PM, recorded every 0.1 ms (shown every 100 image frames = every 10 ms). The colors in the trajectory indicate the diffusion in different plausible compartments (order of appearance: purple, blue, green, orange, red, and then back to purple; the same color order is used throughout this report), detected by the TILD analysis software (Fig. S5). (D) Typical single-molecule trajectories of Cy3-DOPE and Cy3-Tf-TfR in intact and actin-depleted blebbed apical PMs, recorded at normal video rate and enhanced rates. The order of the compartments that the molecule visited (parenthesized integers) and the residency times there, as determined by the TILD analysis, are indicated (intact PMs).

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