Anti-Histone H3 (acetyl K9)抗体[AH3-120] - ChIP Grade (ab12179)

概述

  • 产品名称
    Anti-Histone H3 (acetyl K9)抗体[AH3-120] - ChIP Grade
    参阅全部 Histone H3 一抗
  • 描述
    小鼠单克隆抗体[AH3-120] to Histone H3 (acetyl K9) - ChIP Grade
  • 宿主
    Mouse
  • 经测试应用
    适用于: Flow Cyt, IHC-P, ChIP, WB, ELISA, ICC/IFmore details
  • 种属反应性
    与反应: Mouse, Human, Arabidopsis thaliana, Drosophila melanogaster, Rice
    预测可用于: Rat, Chicken, Cow, Xenopus laevis, Caenorhabditis elegans, a wide range of other species
  • 免疫原

    Synthetic peptide corresponding to Histone H3 aa 7-20 (N terminal) (acetyl K9).
    Sequence:

    ARKSTGGKAPRKQL

  • 阳性对照
    • IHC-P: Human normal colon FFPE tissue sections.
  • 常规说明
    Storage in frost-free freezers is not recommended. If slight turbidity occurs upon prolonged storage, clarify the solution by centrifugation before use. Working dilutions should be discarded if not used within 12 hours.

性能

应用

Our Abpromise guarantee covers the use of ab12179 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
Flow Cyt Use at an assay dependent concentration. PubMed: 19584087

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

IHC-P Use a concentration of 10 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ChIP Use 5-10 µg for 25 µg of chromatin.
WB Use a concentration of 1 - 2 µg/ml. Detects a band of approximately 17 kDa.
ELISA Use at an assay dependent concentration.
ICC/IF 1/1000.

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 H3 family.
  • 发展阶段
    Expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
  • 翻译后修饰
    Acetylation is generally linked to gene activation. Acetylation on Lys-10 (H3K9ac) impairs methylation at Arg-9 (H3R8me2s). Acetylation on Lys-19 (H3K18ac) and Lys-24 (H3K24ac) favors methylation at Arg-18 (H3R17me).
    Citrullination at Arg-9 (H3R8ci) and/or Arg-18 (H3R17ci) by PADI4 impairs methylation and represses transcription.
    Asymmetric dimethylation at Arg-18 (H3R17me2a) by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 (H3R8me2s) by PRMT5 is linked to gene repression. Asymmetric dimethylation at Arg-3 (H3R2me2a) by PRMT6 is linked to gene repression and is mutually exclusive with H3 Lys-5 methylation (H3K4me2 and H3K4me3). H3R2me2a is present at the 3' of genes regardless of their transcription state and is enriched on inactive promoters, while it is absent on active promoters.
    Methylation at Lys-5 (H3K4me), Lys-37 (H3K36me) and Lys-80 (H3K79me) are linked to gene activation. Methylation at Lys-5 (H3K4me) facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 (H3K79me) is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are linked to gene repression. Methylation at Lys-10 (H3K9me) is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 (H3S10ph) and acetylation of H3 and H4. Methylation at Lys-5 (H3K4me) and Lys-80 (H3K79me) require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are enriched in inactive X chromosome chromatin.
    Phosphorylated at Thr-4 (H3T3ph) by GSG2/haspin during prophase and dephosphorylated during anaphase. Phosphorylation at Ser-11 (H3S10ph) by AURKB is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at Ser-11 (H3S10ph) by RPS6KA4 and RPS6KA5 is important during interphase because it enables the transcription of genes following external stimulation, like mitogens, stress, growth factors or UV irradiation and result in the activation of genes, such as c-fos and c-jun. Phosphorylation at Ser-11 (H3S10ph), which is linked to gene activation, prevents methylation at Lys-10 (H3K9me) but facilitates acetylation of H3 and H4. Phosphorylation at Ser-11 (H3S10ph) by AURKB mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. Phosphorylation at Ser-11 (H3S10ph) is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylated at Ser-29 (H3S28ph) by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation. Phosphorylation at Thr-7 (H3T6ph) by PRKCBB is a specific tag for epigenetic transcriptional activation that prevents demethylation of Lys-5 (H3K4me) by LSD1/KDM1A. At centromeres, specifically phosphorylated at Thr-12 (H3T11ph) from prophase to early anaphase, by DAPK3 and PKN1. Phosphorylation at Thr-12 (H3T11ph) by PKN1 is a specific tag for epigenetic transcriptional activation that promotes demethylation of Lys-10 (H3K9me) by KDM4C/JMJD2C. Phosphorylation at Tyr-42 (H3Y41ph) by JAK2 promotes exclusion of CBX5 (HP1 alpha) from chromatin.
    Monoubiquitinated by RAG1 in lymphoid cells, monoubiquitination is required for V(D)J recombination (By similarity). Ubiquitinated by the CUL4-DDB-RBX1 complex in response to ultraviolet irradiation. This may weaken the interaction between histones and DNA and facilitate DNA accessibility to repair proteins.
  • 细胞定位
    Nucleus. Chromosome.
  • Information by UniProt
  • 数据库链接
  • 别名
    • H3 histone family, member A antibody
    • H3/A antibody
    • H31_HUMAN antibody
    • H3FA antibody
    • Hist1h3a antibody
    • HIST1H3B antibody
    • HIST1H3C antibody
    • HIST1H3D antibody
    • HIST1H3E antibody
    • HIST1H3F antibody
    • HIST1H3G antibody
    • HIST1H3H antibody
    • HIST1H3I antibody
    • HIST1H3J antibody
    • histone 1, H3a antibody
    • Histone cluster 1, H3a antibody
    • Histone H3.1 antibody
    • Histone H3/a antibody
    • Histone H3/b antibody
    • Histone H3/c antibody
    • Histone H3/d antibody
    • Histone H3/f antibody
    • Histone H3/h antibody
    • Histone H3/i antibody
    • Histone H3/j antibody
    • Histone H3/k antibody
    • Histone H3/l antibody
    see all

图片

  • Chromatin was prepared from Hela cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The  ChIP was performed with 25µg of chromatin, 5µg of  ab12179 (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). Primers and probes are located in the first kb of the transcribed region.
        

  • IHC image of ab12179 staining Histone H3 (acetyl K9) 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 ab12179, 10µg/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.

  • ab12179 at a 1/1000 dilution staining Stylonychia lemnae (single cell organism, transcriptionally active macronucleus) by ICC/IF. The cells were paraformaldehyde fixed and incubated with the antibody for 12 hours. An Alexa Fluor ® 488 conjugated goat anti-mouse IgG antibody was used as the secondary.

    In the image Histone H3 (acetyl K9) staining is red and is found in macronuclei only. Micronuclei remain unstained and are shown in blue (nucleic acid counterstain). Alpha tubulin is also stained (green).

    See Abreview

  • SK-N-SH cells fixed in 4% paraformaldehyde, permeabilized in 0.5% Triton X-100 and incubated for 1 hour with ab12179 (1/1000 dilution). ab12179 staining is localized to the nucleus (red). The cells were counterstained with DAPI (blue). 100x magnification.

  • All lanes : Anti-Histone H3 (acetyl K9) antibody [AH3-120] - ChIP Grade (ab12179) at 1 µg/ml

    All lanes : Histone fraction isolated from HeLa cells

    Secondary
    All lanes : Goat Anti-Mouse IgG-Peroxidase

文献

This product has been referenced in:
  • Xu J  et al. Super-Resolution Imaging of Higher-Order Chromatin Structures at Different Epigenomic States in Single Mammalian Cells. Cell Rep 24:873-882 (2018). Read more (PubMed: 30044984) »
  • Rodnoi P  et al. Neuropeptide Y expression marks partially differentiated ß cells in mice and humans. JCI Insight 2:N/A (2017). ChIP ; Mouse . Read more (PubMed: 28614797) »
See all 31 Publications for this product

客户评价及客户问答

1-9 of 9 Abreviews or Q&A

Application
IHC - Wholemount
Sample
Zebrafish Embryo (somite)
Specification
somite

Marta Lombó Alonso

Verified customer

提交于 Nov 25 2016

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Skin)
Permeabilization
No
Specification
Skin
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 22°C
Fixative
Paraformaldehyde

Ahmar Aziz

Verified customer

提交于 Jul 08 2016

Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Astatotilapia burtoni Tissue sections (Brain hypothalamus)
Antigen retrieval step
None
Permeabilization
Yes - 0.2% TritonX
Specification
Brain hypothalamus
Blocking step
10% normal serum and 1% BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C
Fixative
Paraformaldehyde

Sebastian Alvarado

Verified customer

提交于 Dec 17 2015

Application
Western blot
Sample
Mouse Tissue lysate - whole (spinal cord, brain, liver)
Gel Running Conditions
Non-reduced Non-Denaturing (Native) (12)
Loading amount
60 µg
Specification
spinal cord, brain, liver
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C

Abcam user community

Verified customer

提交于 Oct 22 2015

Question

I don’t know if I’m using the right lysis method to access nuclear proteins
4) Sample preparation:
Type of sample (whole cell lysates, fraction, recombinant protein…): whole cell lysate from HEK 293T
Lysis buffer PBS with 1% Triton-X-100
Protease inhibitors: n/a
Phosphatase inhibitors n/a
Amount of Lysate IPed ug or number of cells did not do an IP; did a western blot of the lysis
Lysate precleared with matrix : yes/no
Positive control
Negative control

5) IP antibody:
Amount/concentration or dilution used
Conjugation:
Antibody noncovalently bound to matrix before incubation with lysate sample?
Antibody noncovalently bound to matrix after incubation with lysate sample?
Antibody covalently bound to matrix - please describe briefly?
Antibody-lysate incubation time:
Incubation temperature

6) IP Matrix (e. beads):
Type of matrix
Amount of matrix

7) Washing after antibody-lysate incubation:
Buffer
Number of washes

8) IP analysis:
Verification method: e.g reprobe in western blotting

Describe verification method briefly:

OR if Western blotting reprobe performed please indicate the following:

Reducing agent DTT
Boiling for ≥5 min? yes
Protein loaded ug/lane or cells/lane .0043 ug

9) Percentage of gel10 %
Type of membrane nitrocellulose
Protein transfer verified yes, using rainbow marker
Blocking agent and concentration .25% nonfat milk in PBST – (used Millipore Snap ID system)
Blocking time 1 minute
Blocking temperature RT

10) Primary antibody (If more than one was used, describe in “additional notes”) : ab1791

Concentration or dilution 2 ug/ml
Diluent buffer PBST + .25% milk
Incubation time 10 minutes (using Millipore Snap ID system)
Incubation temperature: rt

11) Secondary antibody: goat anti-mouse HRP conjugate from pierce
Species:goat
Reacts against: mouse
Concentration or dilution 1:333
Diluent buffer PBST + .25% milk
Incubation time 10 mins (Used Millipore Snap ID system)
Incubation temperature: RT
Fluorochrome or enzyme conjugate: HRP

12) Washing after primary and secondary antibodies:
Buffer PBST
Number of washes 3x30 mls through Millipore Snap ID

Detection method chemiluminescence

Was the method successfully used to detect protein from non-IPed samples? yes

Document attachment: Attaching images of your blot is strongly recommended and can greatly speed up our investigation of your problem.


This questionnaire doesn’t really even seek to answer the basic question I have, which is: should the use of 1% Triton X-100 in my lysis buffer suffice to lyse the nuclei of HEK cells? Or do I need to do something more?

Read More
Answer

As Histones can be a bit tricky, we usually suggest using a nuclear fraction lysate or to follow our Histone extraction protocol https://www.abcam.com/ps/pdf/protocols/histone%20extraction%20protocol.pdf

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
ChIP
Sample
Rice Cell lysate - whole cell (14 days old seedlings grown under dark condition)
Specification
14 days old seedlings grown under dark condition
Type
Native ChIP (N-ChIP)
Detection step
Real-time PCR
Positive control
Gene of interest
Negative control
No Antibody control

豊 佐藤

Verified customer

提交于 Dec 07 2007

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (MEFs SV40 transformed)
Specification
MEFs SV40 transformed
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.5% triton 10 min RT
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: RT°C

Mr. Mark Rochman

Verified customer

提交于 Aug 13 2007

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Stylonychia lemnae Cell (single cell organism)
Specification
single cell organism
Fixative
Paraformaldehyde
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 4%

Dr. Jan Postberg

Verified customer

提交于 Dec 12 2006

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Arabidopsis thaliana Cell (Landsberg culture cell)
Specification
Landsberg culture cell
Fixative
Paraformaldehyde
Blocking step
BSA as blocking agent for 1 hour(s) and 40 minute(s) · Concentration: 3 %

Dr. Peter McKeown

Verified customer

提交于 Nov 16 2006

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

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