Antibody uptake assay and in vivo imaging to study intracellular trafficking of Notch and Delta in Drosophila.

Notch signaling depends on regulated intracellular trafficking of the receptor and its ligands (Kopan and Ilagan, Cell 137:216-233, 2009; Le Borgne et al., Development 132:1751-1762, 2005). Here we describe two methods to study the intracellular trafficking of Notch and Delta in Drosophila. First, an ex vivo antibody uptake assay is used to monitor endocytosis of Notch and Delta by living cells in dissected explants (Le Borgne and Schweisguth, Dev Cell 5:139-148, 2003). Second, real-time imaging of fluorescent proteins that are expressed at physiological levels is used to study trafficking of Notch in living flies (Venken et al., Science 314:1747-1751, 2006; Couturier et al., Nat Cell Biol 14, 131-139, 2012).