Fluorescence micrographs showing two successive rounds of vesicle fusion (mixing), reaction, and budding/fission (aliquoting).

<p>The 5-µm scale bar in panel (a) also applies to panel (b) and to panels (e)–(h), whereas the 10-µm scale bar in panel (c) also applies to panel (d).</p

Brightfield micrographs showing the fusion, budding, and fission processes of GUVs.

<p>Fission (complete detachment) of daughter GUVs after budding was induced by lowering the temperature close to the <i>T_m</i> of the lipids.</p

Fusion, reaction, and budding of vesicles.

<p>The upper images show GUVs before fusion. The red channel shows the marker for the substrate-containing GUV, whereas the yellow channel shows the marker for the enzyme-containing GUV. The middle images show the budding transformation process after electrofusion. The lower images show daughter GUVs after budding. The increased fluorescence in the green channel indicates the occurrence of the enzymatic reaction.</p

(a) Overview of the experimental setup.

<p>Giant vesicles are manipulated with optical tweezers and fused with an electrical pulse. (b) Schematic of the chemical-handling processes using GUVs. Reagent mixing is induced by fusion, and the reaction products are aliquoted by vesicle division. These processes are repeated for sequential (bio)chemical reactions. In the following experiments, reactions that fluoresce upon reagent mixing are used. The fluorescence in the GUV resulting from the first fusion and reaction is photobleached before the second reaction.</p

Histograms of the M-m ratio for different concentrations of PEG lipid micelles.

<p>(a) 0 <i>μ</i>M, (b) 2.58 <i>μ</i>M, (c) 5.16 <i>μ</i>M, (d) 10.32 <i>μ</i>M, (e) 20.63 <i>μ</i>M, (f) 41.25 <i>μ</i>M, (g) 82.5 <i>μ</i>M, and (h) 165 <i>μ</i>M. Characteristic peaks were indicated as arrows.</p

Histograms of the M-m ratio for different concentrations of PEG lipid micelles.

<p>(a) 0 <i>μ</i>M, (b) 2.58 <i>μ</i>M, (c) 5.16 <i>μ</i>M, (d) 10.32 <i>μ</i>M, (e) 20.63 <i>μ</i>M, (f) 41.25 <i>μ</i>M, (g) 82.5 <i>μ</i>M, and (h) 165 <i>μ</i>M. Characteristic peaks were indicated as arrows.</p

A schematic for vesicle image analysis.

<p>Vesicles are reacted with PEG lipid micelles to induce shape transformations and images are taken by a confocal microscope. Each vesicle image was binarized separately and approximated with an ellipsoid to measure the lengths of major and minor axes.</p

Reconstructed 3D images of a transforming lipid vesicle.

<p>(a) immediately after the addition of PEG lipid micelles, (b) after 2 min, (c) after 4 min (d) after 8 min. Numbers below images indicate the reduced volume <i>v</i>. The concentration of PEG lipid was 2.58 <i>μ</i>M.</p

A shape transformation of lipid vesicle induced by PEG lipid micelles.

<p>(a) A spherical vesicle transformed into (b) a disc-like oblate, (c) cylindrical prolate, and eventually divided into (d) two large and small vesicles.</p

Temporal changes of the mean and variance of the M-m ratio.

<p>Temporal changes of the mean and variance of the M-m ratio.</p