Streptavidin biotin peroxidase complex sabc kit

To examine the spatial distribution of GRP in chicken anterior pituitaries, qPCR, Western blot and immunohistochemical staining (IHC) were used in this experiment, as described in our recent studies (Meng et al. 2014 (link), Mo et al. 2015) . In brief, anterior pituitaries collected from adult female chickens were washed with PBS to remove any contaminating blood cells. The cephalic lobe and caudal lobe were separated carefully by scalpels, and their respective tissue lysates and total RNA were prepared for Western blot detection of cGRP (or cGH/β-actin) protein expression and qPCR assay of cGRP mRNA expression, respectively. For IHC, anterior pituitaries from adult female chickens were fixed in 4% paraformaldehyde, paraffinized and sliced into sections. IHC staining was performed in the pituitary sections using a Streptavidin-Biotin-Peroxidase Complex (SABC) kit (Boster Biological Technology Ltd) according to the manufacturer's instructions. Goat anti-GRP antibody (1:300) was used to probe the spatial distribution of cGRP in chicken pituitaries. Pituitary sections incubated with cGRP 27 -preabsorbed GRP antibody (incubated overnight at 4°C with 100 ng/mL of cGRP 27 ) was used as a negative control in parallel.