Cryostat sections (10 μm) of embryos were processed for anti-nitrotyrosine immunohistochemistry using a specific antibody (Millipore, Billerica, MA, USA; Ref. 06-284, 1:1000 dilution). Immunodetection was carried out with a secondary biotinylated goat anti-rabbit antiserum (Bio-Rad) using immunoperoxidase staining. The expression of nitrotyrosine was scored by quantifying the number of positive cells per region of interest from digital images using NIH ImageJ software. In all sections regions of interest located both on the left and right sides of the embryo were scored independently. Counterstaining was carried out with haematoxylin. Immunofluorescence on pancreas sections with an insulin antibody (Ab7842, Abcam; 1/100 dilution) was performed as described35.