Recombinant
RabMAb

Recombinant Anti-Histone H2A.Z antibody [EPR6171(2)(B)] - ChIP Grade (ab150402)

Overview

  • Product name

    Anti-Histone H2A.Z antibody [EPR6171(2)(B)] - ChIP Grade
    See all Histone H2A.Z primary antibodies
  • Description

    Rabbit monoclonal [EPR6171(2)(B)] to Histone H2A.Z - ChIP Grade
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IHC-P, ICC/IF, Flow Cyt, ChIPmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human Histone H2A.Z aa 1-100. The exact sequence is proprietary.

  • Positive control

    • WB: Neuro-2a, HeLa, HepG2, RAW 264.7, C6 and PC-12 cell lysates. IHC-P: Human colon and lung carcinoma tissues, human breast tissue, mouse and rat cerebrum tissue. ICC/IF: HepG2 and HeLa cells. Flow Cyt: HeLa cells.
  • General notes

     ab150402 is a nuclear marker.

     

     

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab150402 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB 1/1000 - 1/10000. Predicted molecular weight: 13 kDa.
IHC-P 1/2000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

For unpurified format use at 1/250 - 1/500 dilution.

ICC/IF 1/100 - 1/250.
Flow Cyt 1/2500.
ChIP Use 2 µg for 25 µg of chromatin.

Target

  • Function

    Variant histone H2A which replaces conventional H2A in a subset of nucleosomes. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. May be involved in the formation of constitutive heterochromatin. May be required for chromosome segregation during cell division.
  • Sequence similarities

    Belongs to the histone H2A family.
  • Post-translational
    modifications

    Monoubiquitination of Lys-122 gives a specific tag for epigenetic transcriptional repression.
    Acetylated on Lys-5, Lys-8 and Lys-12 during interphase. Acetylation disappears at mitosis.
    Monomethylated on Lys-5 and Lys-8 by SETD6. SETD6 predominantly methylates Lys-8, lys-5 being a possible secondary site.
    Not phosphorylated.
  • Cellular localization

    Nucleus. Chromosome.
  • Information by UniProt
  • Database links

  • Alternative names

    • H2A histone family member Z antibody
    • H2A.z antibody
    • H2A/z antibody
    • H2afz antibody
    • H2AZ antibody
    • H2AZ_HUMAN antibody
    • Histone H2A.Z antibody
    • MGC117173 antibody
    see all

Images

  • All lanes : Anti-Histone H2A.Z antibody [EPR6171(2)(B)] - ChIP Grade (ab150402) at 1/5000 dilution (Purified)

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates
    Lane 3 : Neuro-2a (Mouse neuroblastoma neuroblast) whole cell lysates
    Lane 4 : RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
    Lane 5 : C6 (Rat glial tumor glial cell) whole cell lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 13 kDa
    Observed band size: 13 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human breast tissue sections labeling Histone H2A.Z with purified ab150402 at 1/2000 dilution (1.09 µg/ml). Heat mediated antigen retrieval was performed using heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

  • Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol*. Cells were fixed with formaldehyde for 10 minutes.
    The ChIP was performed with 25 µg of chromatin, 2 µg of ab150402 (red), and 20 µl of Protein A/G sepharose beads. 2 µg of rabbit normal IgG was added to the beads control (gray). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active and inactive loci, Sybr green approach for heterochromatic loci).
    Primers and probes are located in the first kb of the transcribed region.
    *http://www.abcam.com/resources?keywords=X%20ChIP%20protocol

  • Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Histone H2A.Z with purified ab150402 at 1/200 dilution (10 µg/ml). Cells were fixed in 100% Methanol. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

  • Flow cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Histone H2A.Z (red) with purified ab150402 at a 1/2500 dilution. Cells were fixed with 80% methanol and permeabilized with 0.1% Tween-20. A goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730). Blue (unlabeled control) - Cells without incubation with the primary and secondary antibodies.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat cerebrum tissue sections labeling Histone H2A.Z with purified ab150402 at 1:2000 dilution (1.09 µg/ml). Heat mediated antigen retrieval was performed using heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse cerebrum tissue sections labeling Histone H2A.Z with purified ab150402 at 1/2000 dilution (1.09 µg/ml). Heat mediated antigen retrieval was performed using heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

  • Chromatin was prepared from HeLa (Human epithelial cell line from cervix adenocarcinoma) cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab150402 (unpurified) (blue), and 20µl of Anti rabbit IgG sepharose beads. 2μg of rabbit normal IgG was added to the beads control (yellow).  The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach). Primers and probes are located in the first kb of the transcribed region.

  • Immunohistochemical analysis of paraffin-embedded human colon tissue labeling Histone H2A.Z with ab150402 (unpurified) at 1/250 dilution.

  • Immunofluorescent analysis of HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling Histone H2A.Z with ab150402 (unpurified) at 1/100 dilution.

  • All lanes : Anti-Histone H2A.Z antibody [EPR6171(2)(B)] - ChIP Grade (ab150402) at 1/1000 dilution (unpurified)

    Lane 1 : Recombinant Human octamers containing H2A at 1 µg
    Lane 2 : Recombinant Human octamers containing H2A at 0.5 µg
    Lane 3 : Native recombinant octamers K562 cells at 3 µg
    Lane 4 : Native recombinant octamers K562 cells at 1.5 µg
    Lane 5 : Recombinant Human octamers containing H2A.Z.2.1 at 0.5 µg
    Lane 6 : Recombinant Human octamers containing H2A.Z.1 at 0.5 µg

    Secondary
    All lanes : HRP-conjugated donkey anti-rabbit IgG polyclonal at 1/10000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 13 kDa
    Observed band size: 15 kDa
    why is the actual band size different from the predicted?


    Exposure time: 5 minutes

    See Abreview

  • Immunohistochemical analysis of paraffin-embedded human lung carcinoma tissue labeling Histone H2A.Z with ab150402 (unpurified) at 1/250 dilution.

  • Immunohistochemical analysis of paraffin embedded human cervical carcinoma tissue using ab150402 (unpurified) showing positive staining.

  • Immunohistochemical analysis of paraffin embedded normal human kidney tissue using ab150402 (unpurified) showing positive staining.

  • Immunohistochemical analysis of paraffin embedded human prostate hyperplasia tissue using ab150402 showing positive staining.

  • Immunohistochemical analysis of paraffin embedded human ovarian carcinoma tissue using ab150402 (unpurified) showing positive staining.

References

This product has been referenced in:

  • Yang B  et al. H2A.Z regulates tumorigenesis, metastasis and sensitivity to cisplatin in intrahepatic cholangiocarcinoma. Int J Oncol 52:1235-1245 (2018). Read more (PubMed: 29532867) »
  • Cheung P  et al. Single-Cell Chromatin Modification Profiling Reveals Increased Epigenetic Variations with Aging. Cell 173:1385-1397.e14 (2018). Read more (PubMed: 29706550) »
See all 3 Publications for this product

Customer reviews and Q&As

Application
Western blot
Loading amount
0.5 µg
Gel Running Conditions
Reduced Denaturing (4-20% TGX (BioRad))
Sample
Human Recombinant protein (Recombinant octamers & purified K562 cell histones)
Specification
Recombinant octamers & purified K562 cell histones
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 20°C

Dr. Ragnhild Eskeland

Verified customer

Submitted Jan 27 2015

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