Immunofluorescence and in situ hybridization

Immunolabelling was performed as described (Vieira et al., 2007 (link)) using rabbit anti-mouse TUJ1 (1:250; MRB-435P, BioLegend) and mouse anti-rat ISL1 (1:100; 39.4D5, DSHB) followed by Alexa Fluor 594-conjugated rabbit anti-goat Fab (1:200; A21223, Thermo Fisher) and Alexa Fluor 488-conjugated rabbit anti-mouse Fab (1:200; A11017, Thermo Fisher) or horseradish-peroxidase-conjugated goat anti-rabbit antibody (1:200; P0448, DAKO). To detect blood vessels, we used biotinylated isolectin B4 (1:500; L2140, Sigma) followed by Alexa Fluor 633-conjugated streptavidin (1:200; S-21375, Thermo Fisher). We used digoxigenin-labelled RNA probes for Isl1 (Schwarz et al., 2004 (link)), Hoxb1 (Gavalas et al., 2003 (link)), Hs6st1, Hs6st2 (Sedita et al., 2004 (link)), Erm (Trokovic et al., 2005 (link)) and Fgfr1-4 (Pringle et al., 2003 (link); Trokovic et al., 2003 (link)).