Immunostaining of Drosophila Tissues

Tissues were fixed in 3% paraformaldehyde in PBS for 30 min then immunostained as described previously [20 (link)]. Mouse anti-Coracle and anti-Armadillo antibodies (Developmental Studies Hybridoma Bank (DSHB)) and anti-Fibrillarin (Thermo Fisher Scientific) were used at 1:10, 1:400, and 1:400 dilution respectively. Goat anti-mouse IgG labeled with Alexa Fluor 594 (BD Biosciences) was used at 1:500 dilution as the secondary antibody. Guinea pig anti-Lov antibody, prepared and characterized as described previously [20 (link)], was used at a 1:50 dilution followed by goat anti-guinea pig IgG labeled with Alexa Fluor 594 or 488. DNA was stained with DAPI (4’,6-diamidino-2-phenylindole) at 1μg/ml prior to mounting in 70% glycerol. Fat body was simultaneously fixed and stained in 37% formaldehyde containing five units/ml of Oregon-Green 488nm Phalloidin (Molecular Probes) and DAPI at 1μg/ml.