Axiovert 200m microscope

Fat bodies from 96 hours AEL (after egg laying) larvae were dissected in phosphate-buffered saline (PBS) at room temperature, fixed 25–30 min in 4% formaldehyde, washed twice for 10 min in PBS 0.3% Triton (PBST), blocked 30 min (PBST, 5% BSA, 2% FBS, 0.02% NaN₃), incubated with primary antibodies in the blocking buffer overnight, and washed 4 times for 15 min. Secondary antibodies diluted 1:500 in PBST were added for 1 hour and tissues washed 4 times before mounting in Vectashield (Vector Laboratories) containing DAPI. Brains from 5–10 days old adult female flies were dissected and processed as in a previous study^{69 (link)}.
Images for Figure 2c and Extended Data Figure 3d were acquired on Zeiss Axio Zoom V16. Images for Figure 4b, 4c, Extended Data Figure 1c, Extended Data Figure 7f, and Extended Data Figures 8b, 8c were acquired on a Zeiss AxioVert200M microscope with a 63X or 40X oil immersion objective or 10X objective and a Yokogawa CSU-22 spinning disk confocal head with a Borealis modification (Spectral Applied Research/Andor) and a Hamamatsu ORCA-ER CCD camera. The MetaMorph software package (Molecular Devices) was used to control the hardware and image acquisition. The excitation lasers used to capture the images were 405 nm, 488 nm, and 561 nm. Images for Extended Data Figures 6b, 6c were acquired on iPhone XR camera via a binocular microscope.