Anti pc1 3

Tissues were fixed in 4% paraformaldehyde for 30 min at 4°C, embedded in paraffin and 8 µm sections applied to glass slides. These sections were assayed as described previously (Collombat et al., 2003 (link)) using DAPI as counterstain. The primary antibodies used were guinea pig anti-insulin (1/500; DAKO), anti-Pdx1 (1/1,000; provided by C. Wright, Vanderbilt University, Nashville, TN); chicken anti-vimentin (1/4,000; Millipore); rat anti-somatostatin (1/250; Millipore) and anti-BrdU (1/50; Abcam); goat anti-somatostatin (1/500; Santa Cruz); mouse anti-BrdU (1/50; Roche), anti-Ngn3 (1/250; DSHB), and anti-glucagon (1/500; Sigma); rabbit anti-glucagon (1/500; R&D Systems), anti–glut-2 (1/500; Chemicon), anti-PC1/3 (1/500; Millipore), anti-Nkx6.1 (1/3000; NovoNordisk), anti-Pax6 (1/500; BioLegend), anti-Pax4 (1/500; provided by B. Sosa-Pineda, Northwestern University, Evanston, IL), anti-NeuroD1 (1/500; Millipore), and anti-synaptophysin (1/400; Abcam). The secondary antibodies (1/1,000; Invitrogen and Jackson ImmunoResearch) used were Alexa Fluor 488, 594, and 647 and Cy3 and Cy5. For histological analyses, sections were subjected to hematoxylin and eosin or Picro-Sirius red staining. Images were processed as described in Microscope image acquisition.