Figure 4:

Cell-free reconstituted fluorescent sfCherry between two apposing membranes. A) Representative confocal images of reconstituted sfCherry (magenta) that fluoresces in the space between SUVs harboring sCatch-GFP (green) and SUPER templates displaying InterTag-BFP (cyan). Scale bars: 10 μm. B) Box plots comparing the relative sfCherry signal in the presence (magenta) or absence (gray) of SpyCatcher domain. The ratios of sfCherry signal on the SUPER membrane to the background signal were averaged over 30 points along the bead periphery to account for non-uniform distribution of fluorescence over the bead surface. F/F₀ is the mean value of the ratios, averaged across 8 different beads (n=3). The box represents the 25–75th percentiles, and the median is indicated. The whiskers show the minimum and maximum data points. p-values are calculated using two-tailed t-test. C) Representative confocal images of reconstituted sfCherry (magenta) that fluoresces in the space between SUVs harboring sTag-BFP (cyan) and SUPER templates displaying InterCatch-GFP (green). Scale bars: 10 μm. D) Box plots comparing the relative sfCherry signal in the presence (magenta) or absence (gray) of SpyTag domain. The ratios of sfCherry signal on the SUPER membrane to the background signal were averaged over 30 points along the bead periphery to account for non-uniform distribution of fluorescence over the bead surface. F/F₀ is the mean value of the ratios, averaged across 8 different beads (n=3).