Histone H3K9me2 antibody (pAb)

RRID: AB_2793199
Catalog No: 39239 Format: 100 µl $470 Buy
Catalog No: 39240 Format: 10 µl $115 Buy
Catalog No: 39041 Format: 50 µl $275 Buy

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Antibody Type:
Polyclonal
Isotype:
Serum
Purification:
None
Host:
Rabbit
Molecular Weight:
17 kDa
Reactivity:
Human, Mouse, Wide Range Predicted

Applications

ChIP Validated Western Blot Validated Immunofluorescence Validated Dot Blot Validated Immunocytochemistry Validated

Application Notes

Applications Validated by Active Motif:
ChIP: 10 µl per ChIP
ICC/IF: 1:250 - 1:1,000 dilution
WB*: 1:2,000 - 1:5,000 dilution

*Note: many chromatin-bound proteins are not soluble in a low salt nuclear extract and fractionate to the pellet. Therefore, we recommend a High Salt / Sonication Protocol when preparing nuclear extracts for Western blot.

Published Applications

The following applications have been published using this antibody. Unless noted above, Active Motif may not have validated the antibody for use in these applications:

ChIP, ChIP-qPCR
CUT&RUN
Native ChIP
WB
IP
IF

View publications that use Active Motif products & services here. Enter the product number(s) to see publications & applications that use this antibody.

Immunogen

This Histone H3 dimethyl Lys9 antibody was raised against a peptide including dimethyl-lysine 9 of histone H3.

Buffer

Rabbit serum containing 30% glycerol and 0.035% sodium azide. Sodium azide is highly toxic.

 

Histone H3 dimethyl Lys9 antibody tested by ChIP analysis.
Chromatin IP performed using the ChIP-IT® Express Kit (Catalog No. 53008) and HeLa Chromatin (1.5 x 106 cell equivalents per ChIP) using 10 µl of Histone H3 dimethyl Lys9 antibody or the equivalent amount of rabbit IgG as a negative control. Real time, quantitative PCR (RT-qPCR) was performed on DNA purified from each of the ChIP reactions using a primer pair specific for the indicated gene. Data are presented as Fold Enrichment of the ChIP antibody signal versus the negative control IgG using the ddCT method.

Histone H3K9me2 antibody tested by immunofluorescence.
Staining of HeLa cells with Histone H3K9me2 antibody (1:1,000 dilution, top panel) and DAPI (middle panel), and a merge of both images (bottom panel).

Histone H3K9me2 antibody tested by Western blot.
Detection of H3K9me2 by Western blot. The analysis was performed using 20 µg HeLa acid extract and Histone H3K9me2 antibody at a 1:1,000 dilution.

Histone H3K9me2 antibody specificity tested by peptide array analysis.
Peptide array analysis was used to confirm the specificity of this antibody for its intended modification. Histone H3 dimethyl Lys9 antibody was applied at a dilution of 1:15,000 to Active Motif's MODified™ Histone Peptide Array (Catalog No. 13001). The arrays were scanned with ArrayAnalysis Software 7 and the results plotted. Specificity data is shown for the most reactive peptides.

Histone H3K9me2 antibody tested by dot blot analysis.
Dot blot analysis was used to confirm the specificity of Histone H3 dimethyl Lys9 pAb for dimethyl-lysine 9 of histone H3. Decreasing amounts of peptides corresponding to regions around major sites of histone H3 methylation (lysine 4, lysine 9, lysine 27) were spotted onto PVDF and probed with the antibody at a dilution of 1:5,000.
Lane 1: Unmodified Lys4 peptide. Lane 2: Monomethyl Lys4 peptide. Lane 3: Dimethyl Lys4 peptide. Lane 4: Trimethyl Lys4 peptide. Lane 5: Monomethyl Lys9 peptide. Lane 6: Unmodified Lys9 peptide. Lane 7: Dimethyl Lys9 peptide. Lane 8: Trimethyl Lys9 peptide. Lane 9: Unmodified Lys27 peptide. Lane 10: Monomethyl Lys27 peptide. Lane 11: Dimethyl Lys27 peptide. Lane 12: Trimethyl Lys27 peptide.

Background

Histone H3 is one of the core components of the nucleosome. The nucleosome is the smallest subunit of chromatin and consists of 147 base pairs of DNA wrapped around an octamer of core histone proteins (two each of Histone H2A, Histone H2B, Histone H3 and Histone H4). Histone H1 is a linker histone, present at the interface between the nucleosome core and DNA entry/exit points. Histone H1 is responsible for establishing higher-order chromatin structure. Chromatin is subject to a variety of chemical modifications, including post-translational modifications of the histone proteins and the methylation of cytosine residues in the DNA. Reported histone modifications include acetylation, methylation, phosphorylation, ubiquitylation, glycosylation, ADP-ribosylation, carbonylation and SUMOylation; these modifications play a major role in regulating gene expression.

The methylation of histones can occur on two different residues: arginine or lysine. Histone methylation can be associated with transcriptional activation or repression, depending on the methylated residue. Lysine 9 of histone H3 can be mono-, di- or trimethylated by different histone methyltransferases (HMTs) such as SuvH39H1 or G9a. This methylated lysine can be demethylated by histone demethylases as JMJD1A, LSD1 or JMJD2C. Methylation of this residue is mainly associated with transcriptional repression.

Positive Control

Active Motif's HeLa acid extract (Catalog No. 36200) can be used as a positive control for Histone H3 dimethyl Lys9 antibody.

Chromatin IP Assays

This antibody has been validated for use in ChIP and/or ChIP-Seq, and can be used with Active Motif's ChIP-IT® High Sensitivity Kit or our magnetic bead-based ChIP-IT® Express Kits. For an overview of all of our ChIP products, please visit our Chromatin IP Page to learn about ChIP products designed for use with Histone H3K9me2 antibody (pAb).

Storage

Some products may be shipped at room temperature. This will not affect their stability or performance. Avoid repeated freeze/thaw cycles by aliquoting items into single-use fractions for storage at -20°C for up to 2 years. Keep all reagents on ice when not in storage.

Guarantee

This product is guaranteed for 12 months from date of receipt.

This product is for research use only and is not for use in diagnostic procedures.

Application Key

  • ChIP = Chromatin Immunoprecipitation;
  • ChIP = ChIP Sequencing;
  • CUT&RUN = Cleavage Under Targets and Release Using Nuclease;
  • CUT&Tag = Cleavage Under Targets and Tagmentation;
  • DB = Dot Blot;
  • ELISA = Enzyme-linked Immunosorbent Assay;
  • EMSA = Electrophoretic Mobility Shift Assay
  • FC = Flow Cytometry;
  • ICC = Immunocytochemistry;
  • IF = Immunofluorescence;
  • IHC = Immunohistochemistry;
  • IP = Immunoprecipitation;
  • MeDIP = Methyl-DNA Immunoprecipitation;
  • NEU = Neutralizing;
  • WB = Western Blotting