Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade

11 product images

• 

  ChIP - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  Chromatin was prepared from U-2 OS (Human bone osteosarcoma epithelial cell line) 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 ab8580 (blue), and 20 μl 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.

• 

  This image is courtesy of an anonymous customer review

  ChIP - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  
  Chromatin was prepared from human cell lysate - nuclear B cells according to the Abcam X-ChIP protocol.
  

  Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 0.5 μg of ab8580 per μg chromatin in ChIP Buffer fot 16 hours at 4°C. The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region. Negative control: IgG and Gene Desert. Positive control: GAPDH Promoter.

• 

  Western blot - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  All lanes: Western blot - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580) at 1 µg/mL

  All lanes: Calf thymus histone preparation (nuclear lysate) at 0.5 µg

  Secondary

  All lanes: Goat Anti-Rabbit IgG (H+L) HRP- conjugated antibody at 1/50000 dilution

  Performed under reducing conditions.

  Predicted band size: 15 kDa

  Observed band size: 17 kDa

  Exposure time: 8min

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  IHC image of ab8580 staining Histone H3 (tri methyl K4) 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 (pH 6, epitope retrieval solution 1) for 20 minutes. The section was then incubated with ab8580, 1/500 dilution, for 15 minutes 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 (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.

  *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

• 

  This image is courtesy of Ahmad Khalil and Daniel Driscoll, University of Florida College of Medicine.

  Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  Human female lymphoblast immunostained with ab8580 (1:100) (yellowish green) specific for histone H3 lysine 4 (H3-K4) trimethylation; the DNA is stained red with propidium iodide (PI).
  

  Note the inactive X chromosome (arrow) and pericentromeric heterochromatin are largely devoid of this modification.

• 

  Peptide Array - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  All batches of ab8580 are tested in Peptide Array against peptides to different Histone H3 modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H3 - tri methyl K4 peptide (Human Histone H3 (tri methyl K4) peptide ab1342), indicating that this antibody specifically recognises the Histone H3 - tri methyl K4 modification. Slight cross reactivity is observed with the Histone H3 - di methyl K4 modification. Optimization is recommended to avoid array signal saturation.
  

  1. Human Histone H3 (mono methyl K4) peptide ab1340 - Histone H3 - mono methyl K4
  2. Human Histone H3 (tri methyl K4) peptide ab1342 - Histone H3 - tri methyl K4
  3. Human Histone H3 (mono methyl K9) peptide ab1771 - Histone H3 - mono methyl K9
  4. Human Histone H3 (di methyl K9) peptide ab1772 - Histone H3 - di methyl K9
  5. Human Histone H3 (tri methyl K9) peptide ab1773 - Histone H3 - tri methyl K9
  6. Human Histone H3 (mono methyl K27) peptide ab1780 - Histone H3 - mono methyl K27
  7. Human Histone H3 (di methyl K27) peptide ab1781 - Histone H3 - di methyl K27
  8. Human Histone H3 (tri methyl K27) peptide ab1782 - Histone H3 - tri methyl K27
  9. Human Histone H3 (unmodified) peptide ab7228 - Histone H3 - unmodified
  10. Human Histone H3 (di methyl K4) peptide ab7768 - Histone H3 - di methyl K4
• 

  This image is courtesy of a customer review submitted by Dr Eva Bartova

  Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  ab8580 staining cultured human primary fibroblasts by ICC.
  

  Cells were fixed in PFA and permeabilized in Triton X-100 and saponin prior to blocking with 1% BSA for 1 hour at RT. The primary antibody was diluted 1/100 and incubated with the sample for 16 hours at 4°C. An FITC-conjugated rabbit anti-rabbit IgG antibody was used as the secondary.

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  ab8580 staining Histone H3 (tri methyl K4) in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells.
  

  All cells were fixed with 100% methanol (5 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with Anti-Histone H3 (acetyl K27) antibody - ChIP Grade ab4729 at 1/1000 and Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 at 1μg/ml overnight at +4°C, followed by a further incubation at room temperature for 1 hour with goat anti-rabbit Alexa-Fluor^®488 secondary (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) at 2 μg/ml (shown in green) and goat anti-mouse Alexa-Fluor^®594 secondary (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labeled in blue with DAPI.

  Negative controls: 1– Rabbit primary and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no non-specific reaction between primary and secondary antibodies used.

• 

  This image is courtesy of Kirk McManus in the lab of Michael Hendzel, University of Alberta

  Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  Staining (green) with the anti-trimethyl Lysine K4 of Histone H3 antibody (ab8580) shows ringing of regions that appear as nucleoplasmic holes. These represent the positions of splicing factor compartments, which are preferentially associated with active genes and highly acetylated histone H3.

  The antibody, as expected, fails to stain heterochromatin (red).

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  ab8580 staining Histone H3 (tri methyl K4) in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab8580 at 0.1µg/ml and Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour magenta). Nuclear DNA was labelled with DAPI (shown in blue). Also suitable in cells fixed with 4% paraformaldehyde (10 min).Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• 

  This image is courtesy of an anonymous customer review

  ChIP - Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580)

  ChIP was performed with human thyroid cancer cell lysate and 1 µg/µg of ab8580. Lysates were incubated with the primary antibody for 16 hours at 4°C. positive control promoter: GAPDH promoter; positive control enhancer: enhancer region of a published target (10.1093/nar/gkx802); negative control: published negative region (10.1093/nar/gkx802).