Insulin is necessary and sufficient for Clu particle formation. (A-A‴) Stills from live-imaging of starved cluCA06604 germ cells expressing Clu::GFP. Females were raised on standard fly food (no yeast paste) and dissected in Complete Schneider’s. After adding insulin at time zero (A), particles start forming within five minutes (A′). (B) Quantification of the percent single follicles containing Clu particles for females starved on standard food (no yeast paste) or H2O. (C-E″) Clonal analysis in follicle cells. Wild type control clones (FRT82B, GFP−, dotted white line) have Clu particles as expected (C-C″). Clones mutant for TSC1Q87X also have Clu particles (D-D″, dotted white lines). Clones mutant for InR339 lack particles (dotted white lines) (E-E″). (F) Quantification of the number of GFP+ and GFP− clones containing particles for control FRT82B (n = 20), TSC1 (n = 20), and InR339 (n = 11) mutant clones. White = Clu::GFP (A-A‴), GFP (C, D, E), and Clu, (C′, D′, E′), green = Clu, magenta = GFP. For B, error bars = S.D. Scale bars: 20 μm in A‴ for A-A‴, 10 μm in E″ for C-E′.