Supplementary Information
for
Rapid formation of multicellular spheroids in double-emulsion droplets with controllable microenvironment
Hon Fai Chan1, Ying Zhang1, Yi-Ping Ho2, Ya-Ling Chiu1, Youngmee Jung1,3, Kam W. Leong1*
1Department of Biomedical Engineering, Duke University, 101 Science Drive, Durham, NC 27708, USA, 2Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Aarhus C 8000, Denmark, 3 Center for biomaterials, Korea institute of science and technology, 14-gil 5 Hwarangno, Seoungbukgu, Seoul, 136-791, Korea
Supplementary Figure 1. The microfluidics platform adopted for generating DE. The first PDMS chip was used to generate w/o emulsion before they were transferred to the second chip where the w/o/w emulsion were created and collected.
Supplementary Figure 2. Example of diffusion of fluorescent dye from the DE droplets. Fluorescent images showing rhodamine b encapsulated-DE droplets at a) 0 hr, b) 2 hr and c) 4 hr (Scale bar = 100 μm).
Supplementary Figure 3. DE droplets as bioreactor with controlled cell number. a) Plot of experimental data (distribution of cell number in 200 DE droplets) versus Poisson estimation.The diameter of droplet core was 100 μm and the cell density used was 8* 10^6 cells/ml, which gave the average number of cells in each droplet of 4. b) Percentage of viable cell area in spheroids obtained at various time points (n ≥ 10) (Data = mean ± SD).
Supplementary Figure 4. Bright field and live-dead images showing spheroid formation of a) PMEF, b) HepG2 at 2 hr and Caco-2 at 2 hr (c) and 6 hr (d) (Scale bar = 100 μm) .
Supplementary Figure 5. Frequency of hMSC spheroid a) formation and b) retrieval at various cell encapsulation densities (Data = mean ± SD). Phase images showing uniform-sized hMSC spheroids obtained at c) 2* 10^6 cells/ml and d) 10* 10^6 cells/ml (Scale bar = 100 μm).
Supplementary Figure 6. Spheroid composed of hMSC formed in DE stabilized by PEG-PFPE surfactant(Scale bar = 100 μm).
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