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Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, mono methylated at K9.
(Peptide available as ab1771.)
Our Abpromise guarantee covers the use of ab9045 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IP||Use at an assay dependent concentration.|
|WB||1/1000. Detects a band of approximately 15 kDa (predicted molecular weight: 17 kDa).Can be blocked with Human Histone H3 (mono methyl K9) peptide (ab1771). Can be blocked with Histone H3 peptide - mono methyl K9 (ab1771).|
|IHC-P||Use at an assay dependent concentration.|
|Flow Cyt||1/100. Ab171870-Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.|
|ChIP||Use 4-5µg for 106 cells.|
|ICC/IF||Use at an assay dependent concentration.|
ab9045 staining Histone H3 (mono methyl K9) 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 (1/300) 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).
Rabbit polyclonal to Histone H3 K9 Methyl K9 (1/1000)
Peptides at 1 ug/ml
1XTBS, 5%BSA, 0.5% TweenThis antibody shows significantly greater reactivity with mono methyl K9. This can be seen in lane 7, as the addition of ab1771 (mono methyl K9) completely blocks the activity of ab9045. Weaker cross-reactivity is seen against mono methyl K27. This is shown in lane 3, as the addition of ab1780 only partially blocks the activity of ab9045.
Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 min. The ChIP was performed with 25 µg of chromatin, 2 µg of ab9045 (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 for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.
ab9045 staining rat liver tissue sections by IHC-P. Sections were formaldehyde fixed and subjected to heat mediated antigen retrieval in citrate buffer pH 6.0 prior to blocking with 5% serum for 30 minutes at 20°C. The primary antibody was diluted 1/400 and incubated with the sample for 45 minutes at 20°C. A HRP-conjugated goat anti-rabbit antibody was used as the secondary.
Anti-mono methyl lysine 9 of histone H3 (green) has a distribution often associated with euchromatic probes (small foci). Most of these foci localize to regions that contain obvious enrichments of DNA with DAPI staining (red). The perinucleolar chromatin is typically a site enriched in monomethylated lysine 9.
Top left: Mono-methyl Lys 9 (ab9045); Bottom left: DAPI; Top right: Merge of ab9045 (green) and DAPI (red).
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