Anti-Histone H2A抗体- ChIP Grade (ab88770)

概述

  • 产品名称Anti-Histone H2A抗体- ChIP Grade
    参阅全部 Histone H2A 一抗
  • 描述
    兔多克隆抗体to Histone H2A - ChIP Grade
  • 经测试应用适用于: ChIP, IHC-P, ICC/IF, WB, IPmore details
  • 种属反应性
    与反应: Mouse, Cow, Human, Caenorhabditis elegans, Plasmodium falciparum
    预测可用于: Rat, Chicken, Monkey
  • 免疫原

    Synthetic peptide corresponding to Human Histone H2A aa 50 to the C-terminus conjugated to Keyhole Limpet Haemocyanin (KLH).
    (Peptide available as ab100824)

  • 阳性对照
    • This antibody gave a positive signal in the following whole cell lysates: Calf thymus histone; C elegans; HeLa; NIH3T3. IHC-P: FFPE human breast adenocarcinoma tissue sections and human colon.

性能

应用

Our Abpromise guarantee covers the use of ab88770 in the following tested applications.

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

应用 Ab评论 说明
ChIP Use 5 µg for 25 µg of chromatin.
IHC-P Use a concentration of 0.1 - 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ICC/IF Use a concentration of 1 µg/ml.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 17 kDa (predicted molecular weight: 14 kDa).
IP Use a concentration of 5 µg/ml.

靶标

  • 功能Core component of nucleosome. 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.
  • 序列相似性Belongs to the histone H2A family.
  • 翻译后修饰The chromatin-associated form is phosphorylated on Thr-121 during mitosis.
    Deiminated on Arg-4 in granulocytes upon calcium entry.
    Monoubiquitination of Lys-120 by RING1 and RNF2/RING2 complex gives a specific tag for epigenetic transcriptional repression and participates in X chromosome inactivation of female mammals. It is involved in the initiation of both imprinted and random X inactivation. Ubiquitinated H2A is enriched in inactive X chromosome chromatin. Ubiquitination of H2A functions downstream of methylation of 'Lys-27' of histone H3. Monoubiquitination of Lys-120 by RNF2/RING2 can also be induced by ultraviolet and may be involved in DNA repair. Following DNA double-strand breaks (DSBs), it is ubiquitinated through 'Lys-63' linkage of ubiquitin moieties by the E2 ligase UBE2N and the E3 ligases RNF8 and RNF168, leading to the recruitment of repair proteins to sites of DNA damage. Monoubiquitination and ionizing radiation-induced 'Lys-63'-linked ubiquitination are distinct events.
    Phosphorylation on Ser-2 is enhanced during mitosis. Phosphorylation on Ser-2 by RPS6KA5/MSK1 directly represses transcription. Acetylation of H3 inhibits Ser-2 phosphorylation by RPS6KA5/MSK1.
    Symmetric dimethylation on Arg-4 by the PRDM1/PRMT5 complex may play a crucial role in the germ-cell lineage.
  • 细胞定位Nucleus. Chromosome.
  • Information by UniProt
  • 数据库链接
  • 别名
    • H2a 615 antibody
    • H2A antibody
    • H2A GL101 antibody
    • H2A histone family member A antibody
    • H2A.1 antibody
    • H2A.2 antibody
    • H2A/a antibody
    • H2A/m antibody
    • H2A/O antibody
    • H2A/q antibody
    • H2A1B_HUMAN antibody
    • H2AFA antibody
    • H2AFE antibody
    • H2AFL antibody
    • H2AFM antibody
    • H2AFO antibody
    • H2AFQ antibody
    • HIST1H2AE antibody
    • HIST1H2AJ antibody
    • HIST2H2AA antibody
    • HIST2H2AA3 antibody
    • HIST2H2AB antibody
    • HIST2H2AC antibody
    • Histone 1 H2ae antibody
    • Histone 2 H2aa3 antibody
    • Histone 2 H2ab antibody
    • Histone 2 H2ac antibody
    • Histone H2A type 1 B antibody
    • Histone H2A type 1 C antibody
    • Histone H2A type 1 E antibody
    • Histone H2A type 1 J antibody
    • Histone H2A type 1-B/E antibody
    • Histone H2A.2 antibody
    • Histone H2A/a antibody
    • Histone H2A/m antibody
    • MGC74460 antibody
    see all

Anti-Histone H2A antibody - ChIP Grade 图像

  • 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 25µg of chromatin, 5µg of ab88770 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). 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.

  • IHC image of ab88770 staining Histone H2A in human colon formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab88770, 0.1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the negative control (shown on the inset).

     

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Histone H2A was immunoprecipitated using 0.5mg HeLa whole cell extract, 5µg of Rabbit polyclonal to 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, HeLa 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 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab88770.

    Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).

    Band: 14kDa, non specific band - 95kDa: We are unsure as to the identity of this extra band; Histone H2A

  • IHC image of Histone H2A staining in human breast adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab88770, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • All lanes : Anti-Histone H2A antibody - ChIP Grade (ab88770) at 1 µg/ml

    Lane 1 : Calf Thymus Histone Preparation Nuclear Lysate
    Lane 2 : C.elegans Tissue lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 14 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 14 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 2 minutes
  • ICC/IF image of ab88770 stained HeLa cells. The cells were 100% methanol fixed (5 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 ab88770 at 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) at a concentration of 1.43µM. This antibody also gave a positive result in formaldehyde fixed (4%) (10min) HeLa, Hek293, HepG2, and MCF-7 cells, and in Hek293 and HepG2 Methanol (100%) (5min) fixed cells.

  • All lanes : Anti-Histone H2A antibody - ChIP Grade (ab88770) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 14 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 100 kDa,36 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 30 seconds
  • Anti-Histone H2A antibody - ChIP Grade (ab88770) at 1/1000 dilution + Plasmodium falciparum histone extract at 30000000 cells

    Secondary
    HRP-conjugated goat anti-rabbit polyclonal IgG at 1/100000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 14 kDa
    Observed band size : 16 kDa (why is the actual band size different from the predicted?)


    Exposure time : 1 minute

    This image is courtesy of an anonymous Abreview

    See Abreview

Anti-Histone H2A antibody - ChIP Grade (ab88770)参考文献

This product has been referenced in:
  • Kamano S  et al. The Effects of Calcitonin on the Development of and Ca(2+) Levels in Heat-shocked Bovine Preimplantation Embryos In Vitro. J Reprod Dev 60:317-23 (2014). WB ; Cow . Read more (PubMed: 24899099) »

See 1 Publication for this product

Product Wall

Application Western blot
Loading amount 0.371 µg
Gel Running Conditions Reduced Denaturing (20 %)
Sample Xenopus laevis Recombinant protein (Histones and Nucleosome core particle)
Specification Histones and Nucleosome core particle
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

提交于 Mar 30 2015

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Plasmodium falciparum Cell lysate - nuclear (acid extraction of histones from late stage parasi)
Loading amount 3e+007 cells
Specification acid extraction of histones from late stage parasi
Gel Running Conditions Reduced Denaturing (4-12% Bis-Tris)
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 20°C
Username

Abcam user community

Verified customer

提交于 Jan 02 2013

Thank you for contacting Abcam regarding this issue. I have had a chance to look at each of the antibodies to Histone H2A in our catalog and compare the immunogen sequences with your specific needs. Unfortunately none of these products fall exclusiv...

Read More

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"