Agouti Related Protein (AgRP)

Yongheng Cao1, Masanori Nakata, Shiki Okamoto, Eisuke Takano, Toshihiko Yada, Yasuhiko Minokoshi, Yukio Hirata, Kazunori Nakajima, Kristy Iskandar, Yoshitake Hayashi, Wataru Ogawa, Gregory S. Barsh, Hiroshi Hosoda, Kenji Kangawa, Hiroshi Itoh, Tetsuo Noda, Masato Kasuga, Jun Nakae.PDK1-Foxo1 in Agouti-Related Peptide Neurons Regulates Energy Homeostasis by Modulating Food Intake and Energy Expenditure. PLoS ONE 6(4): e18324. doi:10.1371/journal.pone.0018324.

...For immunofluorescence analyses, mice were transcardially perfused with saline followed by 4% paraformaldehyde in 0.1 M phosphate-buffered saline, pH 7.4 (PBS). The brains were dissected and immersed in 4% paraformaldehyde at 4°C overnight and then soaked in 30% sucrose overnight. Frozen, free-floating coronal sections (4 µm thick) were cut through the arcuate nucleus with a microtome (Leica Microsystems). The sections were washed extensively in PBS for 20 min to quench endogenous peroxidase activity. For double staining of PDK1 and AGRP, the sections were stained with a Renaissance Tyramide Signal Amplification kit (#NEL701, Perkin Elmer, Waltham, MA) according to the manufacturer's protocol. The primary antibodies were Ab-241 (Signalway Antibody, Pearland, TX) for PDK1 and GT15023 (Neuromics, Edina, MN) for AGRP; the secondary antibodies were Alexa FluorR 594 chicken anti-rabbit IgG and Alexa FluorR 488 donkey anti-goat IgG (Molecular Probes, Eugene, OR). For double staining of Foxo1 and AGRP, we used an anti-FOXO1A antibody (ab12161, abcamR, Cambridge, UK), respectively. For double staining of FLAG and AGRP, the sections were stained using a Renaissance Tyramide Signal Amplification kit according to the manufacturer's protocol. The primary antibodies were the OctA-Probe (D-8: sc-807, Santa Cruz Biotechnology, Inc, Santa Cruz, CA); the secondary antibodies were Alexa FluorR 594 chicken anti-rabbit IgG and Alexa FluorR 488 donkey anti-goat IgG (Invitrogen, Carlsbad, CA)...

...goat anti-agouti-related protein (AgRP,. 15ug/ml, Neuromics, Bloomington, MN)...

Peter Kobelt, Anna-Sophia Wisser, Andreas Stengel, Miriam Goebel, Tobias Inhoff, Steffen Noetzel, R¨udiger W. Veh, Norbert Bannert, Ivo van der Voort, Bertram Wiedenmann, Burghard F. Klapp, Yvette Tache, Hubert Monnikes. Peripheral injection of ghrelin induces Fos expression in the dorsomedial hypothalamic nucleus in rats doi: 10.1016/j.brainres.2008.01.054

...AgRP; Neuromics, Minnesota, USA; 1:500 in a solution of 5% w/v donkey ....