Microfluidic device design. (a) Schematic showing a trapped GUV isolated by a donut valve. (b) Enlargement showing the fluid flow being diverted around the trap in the presence of a GUV, before the donut is lowered. (c) Bright-field image showing 30 of the 60 micro-chambers with a height of 20 μm. Fluidic isolation by the donuts is demonstrated using blue and red food dye at various dilutions. Scale bar: 500 μm. (d) Schematic cross-section illustrating the washing step and the function of the valves.
Trapping of GUVs. (a) Wide-field fluorescence image of a single GUV trapped hydrodynamically by the posts (black). The surrounding calcein solution (green) is diverted around the trap. Fluid flow lines are indicated by the dashed lines. (b) The fluid was then exchanged for water without removing the calcein filled GUV. (c) Bright-field image of the donut (grey scale) overlaid with a fluorescence image of the GUV. The membrane is stained with DiI (red). Scale bars: 20 μm. (d) 3-D rendering (using confocal micrographs and ImageJ) of the calcein fluorescence showing minimum deformation of the trapped vesicle.
Histograms of GUV sizes. (a) Before being introduced into the device, the GUVs have a wide size distribution with an average diameter of 25.1 ± 23.6 μm (blue) (noff-chip = 483). After flushing through the device, the trapped GUVs have a more narrow size distribution with an average diameter of 14.0 ± 4.7 μm (green) (non-chip 20 μm = 64) for 20 μm high channels and 5.8 ± 1.4 μm (black) (n on-chip 10 μm = 53) for 10 μm high channels. Representative vesicles of 8, 17 and 6 μm diameter are shown in (b), (c), and (d) respectively. Scale bar: 5 μm.
αHL pore formation. (a) Confocal fluorescence images of calcein being released from a GUV with 50 μg/ml αHL. Scale bar: 10 μm. (b) Kinetics of calcein release for 0, 0.5, 2.5, and 50 μg/ml αHL using 4 separate vesicles. The control without αHL showed no calcein release. The number of individual experiments at a given αHL concentration was, n = 3. Error bars are calculated from the standard deviation.
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