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Products:Epigenetics and Nuclear Signaling >> Transcription >> Domain Families >> Developmental Families >> OCT
Overview
Product name
Anti-Oct4 antibody - ChIP Grade
See all Oct4 products (19) ...
Description
Rabbit polyclonal to Oct4 - ChIP Grade
Specificity
IHC: This antibody is not suitable for staining of mouse paraformaldehyde-fixed frozen sections. We have no available data on the performance of the antibody using alternative conditions or target species in IHC. Recognises a band of the expected size in mouse and human. Recognises an additional band in mouse but not in human.
Tested applications
CHIPseq, ICC/IF, Flow Cyt, IHC-Fr, IHC-P, WB, ChIPmore details
Cross reactivity
Reacts with
Mouse, Human, Rhesus monkey
Predicted to work with
Rat
Immunogen
Synthetic peptide conjugated to KLH derived from within residues 300 to the C-terminus of Human Oct4.
(Peptide available as ab20650.)
Positive control
Human ES cell lysate
Properties
Form
Liquid
Storage instructions
Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
Storage buffer
Preservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS, pH 7.4
Concentration
Concentration information loading...
Purity
Immunogen affinity purified
Clonality
Polyclonal
Isotype
IgG
Research areas
Developmental Biology >> Embryogenesis >> Embryonic stem cells >> Surface molecules
Developmental Biology >> Reproduction >> Germ cell markers
Epigenetics and Nuclear Signaling >> ChIP'ing antibodies >> ChIP'ing antibodies
Stem Cells >> Germline Stem Cells >> Embryonic Germ Cells
Stem Cells >> Embryonic Stem Cells >> Intracellular
Epigenetics and Nuclear Signaling >> Transcription >> Domain Families >> Developmental Families >> OCT
Applications
Show applications key
Our Abpromise guarantee covers the use of ab19857 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
CHIPseq: Use at an assay dependent dilution. (PubMed: 20526341)
ICC/IF: Use a concentration of 1 - 5 µg/ml.
Flow Cyt: 1/100
IHC-Fr: 1/200((see Abreview))
IHC-P: 1/250Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
WB: Use a concentration of 1 µg/mlDetects a band of approximately 43 kDa (predicted molecular weight: 39 kDa).Can be blocked with Oct4 peptide (ab20650).
ChIP: Use at an assay dependent dilution. (PubMed: 18490265)
Target
Function
Transcription 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 specificity
Expressed 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 similarities
Belongs to the POU transcription factor family. Class-5 subfamily.
Contains 1 homeobox DNA-binding domain.
Contains 1 POU-specific domain.
Developmental stage
Highly expressed in undifferentiated embryonic stem cells and expression decreases gradually after embryoid body (EB) formation.
Domain
The POU-specific domain mediates interaction with PKM2.
Post-translational
modifications
Sumoylation 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 localization
Nucleus. Expressed in a diffuse and slightly punctuate pattern.
Target information above from: UniProt accessionQ01860
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).
Alternative names
- MGC22487 antibody
- Oct 3 antibody
- Oct 4 antibody
see all
Database links
- Entrez Gene: 5460 Human
- Entrez Gene: 18999 Mouse
- Entrez Gene: 294562 Rat
- Omim: 164177 Human
- SwissProt: Q01860 Human
- SwissProt: P20263 Mouse
- Unigene: 249184 Human
- Unigene: 632482 Human
see all
Anti-Oct4 antibody - ChIP Grade images:
Immunofluorescence - Oct4 antibody - Embryonic Stem Cell Marker (ab19857)
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 Oct4-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.
Ludovic Vallier, University of Cambridge
Western blot
All lanes : Anti-Oct4 antibody - ChIP Grade (ab19857) at 1 µg/ml
Lane 1 : MEL-1 (Human embryonic stem cell, male cell line) Whole Cell Lysate (ab27198)
Lane 2 : MEL-2 (Human embryonic stem cell, female cell line) Whole Cell Lysate (ab27196)
Lane 3 : IOUD2 (Mouse embryonic stem cell, selected for Oct4 expression cell line) Whole Cell Lysate (ab27202)
Lane 4 : F9 (Mouse embryonic carcinoma cell line) Whole Cell Lysate (ab27193)
Lane 5 : MEL-1 (Human embryonic stem cell, male cell line) Whole Cell Lysate (ab27198) with Oct4 peptide (ab20650) at 1 µg/ml
Lane 6 : MEL-2 (Human embryonic stem cell, female cell line) Whole Cell Lysate (ab27196) with Oct4 peptide (ab20650) at 1 µg/ml
Lane 7 : IOUD2 (Mouse embryonic stem cell, selected for Oct4 expression cell line) Whole Cell Lysate (ab27202) with Oct4 peptide (ab20650) at 1 µg/ml
Lane 8 : F9 (Mouse embryonic carcinoma cell line) Whole Cell Lysate (ab27193) with Oct4 peptide (ab20650) at 1 µg/ml
Lysates/proteins at 10 µl per lane.
Observed band size : 48 kDa (why is the actual band size different from the predicted?)
Additional bands at : 65 kDa. We are unsure as to the identity of these extra bands.
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.
Immunocytochemistry/ Immunofluorescence - Oct4 antibody - Embryonic Stem Cell Marker (ab19857)
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 1/1000 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).
Immunocytochemistry/ Immunofluorescence - Oct4 antibody - Embryonic Stem Cell Marker (ab19857)
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.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Oct4 antibody - Embryonic Stem Cell Marker (ab19857)
Mouse E6 - E6.5 embryos were sectioned and then fixed in paraformaldehyde. Antigen retrieval was performed using citric acid and the sections permeablized and blocked for 1 hour in serum. Embryos were stained with ab19857 (1/2500) in 10% serum, 1% ovalbumin in PBSTween for 14 hours at 4C. They were then washed and stained with a goat anti-rabbit biotin conjugated antibody (1/2000) and developed using ABC and then DAB for color. Staining is within the epiblast as expected.
Western blot
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 Oct4 peptide (ab20650) at 1 µg/ml
Lane 5 : Mouse embryonic stem cell lysate with Oct4 peptide (ab20650) at 1 µg/ml
Lane 6 : Mouse embryonic germ cell lysate with Oct4 peptide (ab20650) at 1 µg/ml
Lysates/proteins at 20 µg per lane.
Predicted band size : 39 kDa
Observed band size : 43 kDa (why is the actual band size different from the predicted?)
Additional bands at : 55 kDa (possible glycosylated form,cross reactivity).
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:
- Griffith GJet al. Yin-yang1 is required in the mammalian oocyte for follicle expansion. Biol Reprod 84:654-63 (2011).Read more (PubMed: 21123818) »
- Casalino Let al. Control of embryonic stem cell metastability by L-proline catabolism. J Mol Cell Biol 3:108-22 (2011).Read more (PubMed: 21307025) »
See all 41 publications for this product
Publishing research using ab19857? Please let us know so that we can cite the reference in this datasheet
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"