Anti gfp fitc

Wing discs were dissected from wandering third instar larve in Grace's medium (source) and incubated for 1 h to incorporate a BrDU analog, EdU, using the Click-iT Kit (Invitrogen). Discs were fixed in 3.5% formaldehyde in PBS, pH 7.2, permeabilized with 0.1% Triton, blocked with 3% BSA, and labeled. The presence of cleaved caspase was detected using Dcp-1 Antibody (Cell Signaling, #9578) at 1/500 dilution, and detected with anti-rabbit Alexa-647 at 1/750. Anti-GFP-FITC (Rockland, #600-402-215) was used at 1/200 dilution and Hoechst 33344 (Life Technologies) at 1/10,000. Discs were mounted in 4% n-propyl gallate in glycerol and imaged on a Zeiss 510 Meta Confocal Microscope. Images were processed by Adobe Photoshop. Total wing disc and the engrailed domain (GFP positive) were defined with the ‘magnetic lasso’ tool. For the measurements shown in Fig. S1E, to reduce background noise, only the top-most epithelial layer corresponding to the wing pouch was measured. For statistics, measurements within genotypes were checked for normality using the shapiro.test() in R. Differences between genotypes were calculated using t.test() in R. All tests used default options.