Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade

• Supplier: Abcam
• Catalog: ab9050
• Prices: $404.00
• Sizes: 100 µg
• Host: Rabbit
• Clonality: Polyclonal
• Isotype: IgG
• Applications: ICC/IF ICC/IF ChIPseq WB ChIP ChIP ChIP IHC-P
• Species Reactivities: Mouse, Rat, Bovine, Human, Yeast, Xenopus, Arabidopsis thaliana, C. elegans, Drosophila, S. pombe, Zebrafish, Rice, Plant
• Antigen: Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, tri methylated at K36
• Blocking Peptide: Human Histone H3 (tri methyl K36) peptide
• Description: Rabbit Polyclonal
• Gene: HIST3H3
• Conjugate: Unconjugated

Product images

ab9050 staining Histone H3 (tri-methyl K36) in HeLa cells. The cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab9050 at 0.1µg/ml and ab7291 (anti beta-Tubulin) at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat anti -rabbit AlexaFluor®488 secondary (ab150081) at 2 μg/ml (shown in green) and a goat anti-mouse AlexaFluor®594 (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.Negative controls: 1– Rabbit primary antibody and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.

ab9050 staining Histone H3 (tri methyl K36) in Human HEK293 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with Triton X-100 and blocked with 1% serum for 10 minutes at 21°C. Samples were incubated with primary antibody (1/500) for 10 hours at 21°C. A FITC-conjugated goat anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody.See Abreview

ab9050 staining Histone H3 (tri methyl K36) in human differentiated haematopoietic stem cells by Flow Cytometry. Cells were fixed with paraformaldehyde and permeabilized with permeabilization buffer. The sample was incubated with the primary antibody (1/400) for 12 hours at 4°C. An Alexa Fluor® 488-conjugated goat polyclonal anti-rabbit IgG (1/1000) was used as the secondary antibody.Gating Strategy: Isotype negative control (white). See Abreview

Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The  ChIP was performed with 25µg of chromatin, 2µg of ab9050 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified on the GAPDH (active) and MYO-D (inactive) promoters and over the ý-Actin gene (active). Schematic diagram of the ý-Actin gene is shown on the top of the figure. Black boxes represent exons and thin lines represent introns. PCR products are depicted as bars under the gene.

Staining of human tonsil using ab9050 is shown. Antigen retrieval was performed using Tris EDTA at pH9. Nuclei of lymphoid cells in the interfollicular area of a human tonsil stained strongly positive as well as the endothelial cells of blood vessels.

ICC/IF image of ab9050 stained human HeLa cells. The cells were methanol fixed (5 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab9050, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. 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). DAPI was used to stain the cell nuclei (blue).

ab9050 staining rat liver tissue sections by IHC-P.  The section was formaldehyde fixed and subjected to heat mediated antigen retrieval in pH 6.0 citrate buffer prior to being blocked with 5% serum for 30 minutes at 45°C.  The primary antibody was diluted 1/500 and incubated for 45 minutes at 20°C.  A HRP conjugated goat anti-rabbit was used as the secondarySee Abreview

ab9050 staining mouse kidney tissue sections by IHC-P.  The section was formaldehyde fixed and subjected to heat mediated antigen retrieval in pH 6.0 citrate buffer prior to being blocked with 5% serum for 30 minutes at 20°C.  The primary antibody was diluted 1/500 and incubated for 45 minutes at 20°C.  A HRP conjugated goat anti-rabbit was used as the secondarySee Abreview

ChIP using ab9050 at the Pho4 locus in S. pombe. Chromatin extract from S.pombe cells was incubated with 5 ug of ab9050 overnight, and then incubated with Protein A beads for 1 hour. The immunoprecipitated DNA was quantified at the Pho4 locus by RT-PCR. No antibody was added to the beads control (red bars). A schematic diagram of the pho4 gene is shown below the graph. The grey box represents the open reading frame and PCR products are depicted by horizontal arrows. Please see anonymous abreview submitted on 29 October 2008 for additional details.See Abreview

All lanes : Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050) at 1 µg/mlLane 1 : Calf Thymus Histone Preparation Nuclear Lysate Lane 2 : Calf Thymus Histone Preparation Nuclear Lysate with Histone H3 peptide - unmodified K36 at 0.5 µg/mlLane 3 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (mono methyl K36) peptide (ab1783) at 0.5 µg/mlLane 4 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (di methyl K36) peptide (ab1784) at 0.5 µg/mlLane 5 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (tri methyl K36) peptide (ab1785) at 0.5 µg/mlLane 6 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (tri methyl K37) peptide (ab24417) at 0.5 µg/mlLysates/proteins at 0.5 µg per lane.SecondaryGoat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilutionPerformed under reducing conditions.

ab9050 staining Histone H3 (tri methyl K36) in Human Saos-2 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.25% Triton in PBS and blocked with 1% BSA for 1 hour at room temperature. Samples were incubated with primary antibody (1/1000) for 1 hour. An Alexa Fluor®488-conjugated Goat anti-rabbit IgG polyclonal (1/250) was used as the secondary antibody.See Abreview

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