Left: confocal microscopy images of de-mixed droplets in vitro formed by mixing HERD-2.2-GFP plus TC-HERD-2.2-TnaA (top), and HERD-2.2-GFP plus TC-HERD-2.2-FMO (bottom), with fluorescent reporters GFP (green) and TC-ReAsH at 561 nm (red). Scale bar, 5 m. Conditions: 125 mM NaCl, 4% PEG 3350, 20 mM Tris pH 7.5, 500 M HERD-2.2-GFP, 25 M HERD-2.2-TnaA or HERD-2.2-FMO. Right: quantification of the fluorescence intensity (561 nm for TC-ReAsH) in de-mixed droplets formed by HERD-2.2-GFP. Fluorescence intensity was normalized by subtracting the fluorescence intensity within HERD-2.2-GFP droplets containing ReAsH dye but no TC-tagged proteins. Data are represented as mean the standard error from n = 12 (TC-HERD-2.2-TnaA, TC-HERD-2.2-FMO) or n = 18 (TC-HERD-4.1-FMO) independent biological measurements. P = 0.001 (***) by one-way analysis of variance (ANOVA) and Tukey’s honestly significant difference (HSD) post-hoc test.
