Chicken antibody to gfp

Tumors were sliced in half along a plane that aligned perpendicular to the approximate injection site. Following fixation with 4% paraformaldehyde (PFA), samples were embedded in paraffin and sectioned with a Micron HM325 microtome (Walldorf, Germany). Slides containing 5 μm thick sections were deparaffinized and rehydrated through a series of ethanol (EtOH) /water washes. Following a 95°C 25-minute antigen retrieval in Trilogy unmasking solution (Cell Marque, Rocklin, CA, USA). Slides were serum treated for one hour at room temperature with 2% normal goat serum (Vector Laboratories, Burlingame, CA, USA). Chicken antibody to GFP (Abcam, Cambridge, MA, USA) was applied at 1:750 overnight at 4°C. Positive staining was detected with a one-hour incubation in 1:500 Alexafluor goat anti-chicken 488 (Invitrogen, Carlsbad, CA, USA) mounted in Vectashield (Vector Laboratories) with 4’, 6-diamidino-2-phenylindole (DAPI) as counterstain.
Immunohistochemical staining was performed using a Rabbit anti-Calcitonin (1:250; Abcam) antibody as previously described [36 (link)]. Images were captured on a Zeiss Axio Vert.A1 inverted light microscope (Zeiss, Thornwood, NY, USA).

Embryos and pups were fixed for 5 to 8 hr in phosphate-buffered saline (PBS) containing 4% PFA (wt/vol), incubated overnight at 4°C with 20% sucrose in PBS (wt/vol), embedded in OCT compound (Sakura Finetek, Tokyo, Japan), and sectioned with a cryostat to obtain 14-µm-thick coronal sections. For detection of Rorb and BrdU, antigen retrieval was performed by autoclave treatment of the sections for 5 min at 105°C in 0.01 M sodium citrate buffer (pH 6.0). As primary antibodies, we used mouse antibody to BrdU (BD Biosciences, San Diego, CA), rabbit antibody to GFP (MBL, Nagoya, Japan), chicken antibody to GFP (Abcam, Cambridge, UK), rabbit antibody to Rorb (Diagenode, Leige, Belgium), mouse antibody to Rorb (Perseus Proteomics, Tokyo, Japan), goat antibody to Lhx2 (Santa Cruz, Santa Cruz, CA), mouse antibody to Satb2 (Abcam), goat antibody to KCNH5 (Santa Cruz), goat antibody to NetrinG1 (R&D systems, Minneapolis, MN) and rabbit antibody to Tbr1 (kind gift from R. Hevner, University of Washington). Immune complexes were detected with FITC–, TRITC– or Cy5–conjugated secondary antibodies (Jackson ImmunoResearch Laboratories, West Grove, PA). For nuclear staining, we used 1 µg/ml propidium iodide (PI; Molecular Probes, Eugene, OR) and 2 µg/ml DAPI (Molecular Probes). Images were acquired using confocal microscopes (FV300 and FV1000; Olympus, Tokyo, Japan).