Nanog antibody (pAb) tested by ChIP-Seq. ChIP was performed using the ChIP-IT High Sensitivity Kit (Cat. No. 53040) with 30 ug of chromatin from undifferentiated hESC cells and 3 ul of antibody. ChIP DNA was sequenced on the Illumina HiSeq and 9 million sequence tags were mapped to identify Nanog binding sites. The image shows binding across a region of chromosome 12. You can view the complete data set in the UCSC Genome Browser, starting at this specific location, here.
Nanog antibody (pAb) tested by Immunofluorescence Mouse embryonic stem cells (mESCs) grown on mouse embryonic fibroblast feeder cells (MEFs) were fixed with 4% paraformaldehyde for 10 minutes at room temperature. Cells were then permeabilized and blocked by incubating with Blocking Solution containing 5% serum/0.1% Triton X-100 in D-PBS for 2 hours at room temperature. Cells were then incubated with Nanog antibody (Catalog No. 61419, red) at 1:200 dilution overnight at 4°C, washed with D-PBS, and incubated for 2 hours at room temperature with goat anti-mouse Cy3 secondary antibody at 1:250 dilution. Cells were visualized using a Zeiss fluorescent microscope at 20X magnification. Images show that Nanog antibody specifically stains mESC colonies and does not stain MEFs. Absence of Nanog staining in a subset of cells within the colonies suggests differentiation. Scale bars, 100 μm.
Nanog (pAb) tested by Western blot. 20 ug of mouse Embryonic Stem Cell (ESC) extract was run on SDS-PAGE and probed with antibody at 2 ug/ml.