Nile red stain

Microfluidic device fabrication was done by using a polydimethylsiloxane (PDMS) preparation set (Sylgard 184, Dow-corning, United States). The oil-soluble surfactant, polyglycerol polyricinoleate (PGPR) was obtained from Danisco (Denmark). Mineral oil, sodium chloride (NaCl) were from Fischer Scientific (United Kingdom) while Poly(allylamine hydrochloride) (M_w ≈ 17 500), poly(sodium 4-styrenesulfonate) (M_w ≈ 70 000) and water-soluble surfactant, polysorbate 80 (Tween 80) were purchased from Sigma Aldrich (United Kingdom). For bacterial study, the material used were nutrient agar, Luria Bertani broth (LB broth), tryptone, yeast extract and phosphate buffer saline (PBS) all by Oxoid Ltd. (United Kingdom). d(+)-glucose was purchased from Acros Organics (United Kingdom) and Nile red stain was purchased from Invitrogen™ (United Kingdom). Escherichia coli strain SCC1 (MG1655-GFP mutation) expressing green fluorescent protein (E. coli-GFP) stock culture was obtained from Biochemical Engineering Laboratory, University of Birmingham, United Kingdom.

For fat body staining, live flies were anesthetized on ice and dissected in ice-cold PBS. The ventral cuticle was cut away from the abdomen using microscissors, and the internal organs and genitalia were removed, leaving the intact abdominal fat body attached to the dorsal cuticle. Samples were fixed in 4% paraformaldehyde on ice for 30 min followed by 3x 10 min washes in PBS. Tissues were then incubated in Nile Red stain (10 μg/mL; Invitrogen N1142), followed by 3x 10 min washes in PBS, and finally mounted onto a slide in VectaShield with DAPI. Images were obtained by confocal microscopy (Leica SP5 II) at excitation/emission 552/636 nm. Brightness was adjusted in Adobe Photoshop to allow visualization of lipid droplet size.