“…The immunocytochemical analysis of embryonic and postnatal mouse brain tissue was undertaken using protocols published previously Simonian and Herbison, 2001b), using antibodies raised against GnRH [1:5000 -1:20,000, LR1; a gift from R. Benoit (Montreal Hospital, Montreal, Quebec, Canada); or 1:10,000, catalog #20075; DiaSorin (Stillwater, MN)], tyrosine hydroxylase (1:4000, MAB318; Chemicon, Temecula, CA), somatostatin (1:3000; a gift from F. Vandesande, Zoological Institute, Leuven, Belgium), choline acetyltransferase (1:5000, AB144; Chemicon), peripherin (1:4000, AB1530; Chemicon), polysialated neural cell adhesion molecule (PSA-NCAM) (1:1000; a gift from G. Rougon, Institute de Biologie du Development, Marseille, France), and the extracellular domain of EphA5 (1:100, E3654; Sigma-Aldrich, Poole, UK). EphA5 immunofluorescence was undertaken by incubating in the goat affinity-purified EphA5 antibody, followed by biotinylated anti-goat IgGs (Vector Laboratories, Peterborough, UK), Vector Elite ABC reagent, and tyramide-Alexa 594 (Molecular Probes, Eugene, OR).…”