Fixation and Immunostaining of Drosophila Embryos

Drosophila embryos were formaldehyde-fixed and processed as described previously (Tepass et al., 1990 (link)). For Arm staining, embryos were heat-fixed (Gamblin et al., 2014 (link)). Primary antibodies used: mouse anti-2A12 (1:10 dilution; Developmental Studies Hybridoma Bank, DSHB), mouse anti-Cora (1:500; clones C566.9 and C615.16, DHSB), mouse and anti-Tango (1:10; DHSB), mouse anti-Arm (1:250; N2-7A1, DHSB), mouse anti-Fas3 (1:10; 7G10, DHSB), rabbit anti-Verm (1:500; Luschnig et al., 2006 (link)), rabbit anti-Serp (1:500; Luschnig et al., 2006 (link)), rabbit anti-aPKC (1:250; C-20, Santa Cruz Biotechnology) and rat anti-Crb (1:500; this study). Secondary antibodies were conjugated to Cy3 (Jackson Immunoresearch Laboratories) or Alexa Fluor 488 (Molecular Probes). Rhodamine-conjugated Chitin-Binding Probe (New England BioLabs) was used at a concentration of 4 µg/ml, and co-incubated with secondary antibodies.