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The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
Use at an assay dependent concentration. PubMed: 20526341
Use a concentration of 1 - 5 µg/ml.
ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.
Use a concentration of 1 µg/ml. Detects a band of approximately 43 kDa (predicted molecular weight: 39 kDa).
Use at an assay dependent concentration. PubMed: 18490265
FunctionTranscription factor that binds to the octamer motif (5'-ATTTGCAT-3'). Forms a trimeric complex with SOX2 on DNA and controls the expression of a number of genes involved in embryonic development such as YES1, FGF4, UTF1 and ZFP206. Critical for early embryogenesis and for embryonic stem cell pluripotency.
Tissue specificityExpressed in developing brain. Highest levels found in specific cell layers of the cortex, the olfactory bulb, the hippocampus and the cerebellum. Low levels of expression in adult tissues.
Sequence similaritiesBelongs to the POU transcription factor family. Class-5 subfamily. Contains 1 homeobox DNA-binding domain. Contains 1 POU-specific domain.
Developmental stageHighly expressed in undifferentiated embryonic stem cells and expression decreases gradually after embryoid body (EB) formation.
DomainThe POU-specific domain mediates interaction with PKM2.
Post-translational modificationsSumoylation enhances the protein stability, DNA binding and transactivation activity. Sumoylation is required for enhanced YES1 expression. Ubiquitinated; undergoes 'Lys-63'-linked polyubiquitination by WWP2 leading to proteasomal degradation.
Cellular localizationNucleus. Expressed in a diffuse and slightly punctuate pattern.
POU domain class 5 transcription factor 1 antibody
POU domain transcription factor OCT4 antibody
POU domain, class 5, transcription factor 1 antibody
POU-type homeodomain-containing DNA-binding protein antibody
Anti-Oct4 antibody - ChIP Grade images
Immunocytochemistry/ Immunofluorescence - Anti-Oct4 antibody - ChIP Grade (ab19857)This image is courtesy of Ludovic Vallier, University of Cambridge
The image shows a colony of differentiating Human Embryonic Stem Cells double-stained with anti-Oct4 antibody ab19857 (green) and Sox17 antibody (red). The nuclei of Oct-4-positive undifferentiated hESCs stained bright green. Staining was restricted to the nuclei. Oct4-positive nuclei were Sox17-negative, and vice versa. This antibody can be used as a marker of Oct4-positive undifferentiated Human Embryonic Stem Cells.
Flow Cytometry - Anti-Oct4 antibody - ChIP Grade (ab19857)This image is taken from an abreview by Kate Hawkins
Overlay histogram of Oct4 staining in mouse embryonic stem cells using ab19857 at 1:100 dilution. Purple histogram represents negative control (rabbit IgG); green line represents anti-Oct4 antibody (ab19857).
MEL-1 and MEL-2 Human Embryonic Stem Cell Lysates (ab27198 and ab27196), IOUD2 Oct4-expressing Mouse Embryonic Stem Cell Lysate (ab272020) and F9 Mouse Embryonic Carcinoma Cell Lysate (ab27193) all contain Oct4 protein. As expected, a band corresponding to Oct4 was detected in all four of the lysates by Westen Blot using anti-Oct4 antibody ab19857. Each of the Oct4 bands was specifically blocked using the immunising peptide of ab19857. The origin of the additional 65 kDa band detected in Lane 3 is unknown and may be a non-specific band that is recognised, in addition to Oct4, by ab19857 in mouse embryonic stem cells.
Oct4 was immunoprecipitated using 0.5mg E14Tg2a whole cell extract, 5µg of Rabbit polyclonal to Oct4 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, E14Tg2a whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation. Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab19857. Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697). Band: 48kDa: Oct4.
ICC/IF image of ab19857 stained Mouse Embryonic Stem cells. The cells were PFA fixed (4% PFA, 20 min) and incubated with the antibody (ab19857, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue).
The large nuclei of the Feeder cells can be seen in the image; ab19857 does not localise to these nuclei. ab19857 can be seen localising to the much smaller nuclei of the Mouse Embryonic Stem cells.
ICC/IF image of ab19857 stained mouse embryonic stem cells. The cells were 4% formalin fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab19857, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1:00 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1:200 dilution for 1h. DAPI was used to stain the cell nuclei (blue).
Western blot - Anti-Oct4 antibody - ChIP Grade (ab19857)
All lanes : Anti-Oct4 antibody - ChIP Grade (ab19857) at 1 µg/ml
Lane 1 : Human embryonic stem cell lysate Lane 2 : Mouse embryonic stem cell lysate Lane 3 : Mouse embryonic germ cell lysate Lane 4 : Human embryonic stem cell lysate with Human Oct4 peptide (ab20650) Lane 5 : Mouse embryonic stem cell lysate with Human Oct4 peptide (ab20650) Lane 6 : Mouse embryonic germ cell lysate with Human Oct4 peptide (ab20650)
Anti-Oct4 antibody ab19857 only detected a band corresponding to the expected size of Oct4 in human ES cell lysate. In mouse ES and EG cell lysates, ab19857 detected a band of approximately 55 kDa in addition to the expected 39 kDa Oct4 band.
References for Anti-Oct4 antibody - ChIP Grade (ab19857)
This product has been referenced in:
Li Y et al. Abnormal Neural Progenitor Cells Differentiated from Induced Pluripotent Stem Cells Partially Mimicked Development of TSC2 Neurological Abnormalities. Stem Cell Reports8:883-893 (2017).
Read more (PubMed: 28344003) »
Xun J et al. JMJD3 suppresses stem cell-like characteristics in breast cancer cells by downregulation of Oct4 independently of its demethylase activity. Oncotarget8:21918-21929 (2017).
Read more (PubMed: 28423536) »