Histone H3K27me3 antibody (pAb)

RRID: AB_2561020
Catalog No: 39155 Format: 100 µg $455 Buy
Catalog No: 39055 Format: 50 µg $265 Buy
Catalog No: 39157 Format: 10 µg $110 Buy

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Antibody Type:
Polyclonal
Isotype:
IgG
Purification:
Protein A Chromatography
Host:
Rabbit
Molecular Weight:
17 kDa
Reactivity:
Human, Mouse, Wide Range Predicted

Applications

CUT&Tag Validated ChIP Validated ChIP-Seq Validated Western Blot Validated Immunofluorescence Validated Dot Blot Validated Immunohistochemistry Validated Immunocytochemistry Validated

Application Notes

Applications Validated by Active Motif:
ChIP: 5 - 10 µg per ChIP
ChIP-Seq: 5 µg each
ICC/IF: 2 µg/ml dilution
IHC(FFPE): 2 µg/ml dilution
WB*: 0.5 - 2 µg/ml dilution
CUT&Tag: 1 µg per 50 µl reaction*

*This antibody has been validated for CUT&Tag using Active Motif's CUT&Tag-IT™ Assay Kit, Catalog No. 53160.

*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 WB.

For H3K27me3, we also offer AbFlex® H3K27me3 Recombinant Antibody (rAb). For details, see Catalog No. 91167.

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-Seq
Native ChIP
WB
ICC/IF
IHC(P), IHC(F)
ELISA
RNA FISH
Proximity Ligation Assay (PLA)

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 trimethyl Lys27 antibody was raised against a peptide including trimethyl-lysine 27 of histone H3. testing

Buffer

Purified IgG in PBS (pH 7.5) with 30% glycerol and 0.035% sodium azide. Sodium azide is highly toxic. For your convenience, an unpurified serum version (Catalog No. 39156) of this antibody is also available.

References

Histone H3K27me3 antibody (pAb) - 100 µg (Cat. No. 39155)

 
 
 

Histone H3K27me3 antibody (pAb) - 10 µg (Cat. No. 39157)

 
 
 

 

Histone H3 trimethyl Lys27 pAb tested by ChIP-Seq.
ChIP was performed using chromatin from the human iPS 19.11 cell line. ChIP DNA was sequenced on the Illumina GA II and sequence tags were mapped to identify H3K27me3 binding. The image shows H3K27me3 binding across a 140,000 bp region of the HOX gene cluster on chromosome 12.

Histone H3 trimethyl Lys27 antibody tested by immunofluorescence.
HeLa cells stained with Histone H3 trimethyl Lys27 antibody (1.7 µg/ml dilution).
    Top panel: Histone H3 trimethyl Lys27 antibody.
    Middle: DAPI.
    Bottom: Merge of both images.

Histone H3 trimethyl Lys27 antibody tested by Western blot.
HeLa acid extract (20 µg) probed with Histone H3 trimethyl Lys27 antibody (1 µg/ml).

Histone H3 trimethyl Lys27 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 trimethyl Lys27 antibody was applied at a dilution of 1:5,000 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 and those related to the immunogen. Recognition of the H3 trimethyl Lys27 peptides by the antibody blocked by phosphorylation at Ser28.
Array Data File

Histone H3 trimethyl Lys27 antibody tested by dot blot analysis.
Dot blot analysis was used to confirm the specificity of Histone H3 trimethyl Lys27 antibody for trimethyl-lysine 27 of histone H3. 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 µg/ml. The amount of peptide (in picomoles) spotted is indicated next to each row. Lane 1: Unmodified Lys4. Lane 2: Monomethyl Lys4. Lane 3: Dimethyl Lys4. Lane 4: Trimethyl Lys4. Lane 5: Monomethyl Lys9. Lane 6: Unmodified Lys9. Lane 7: Dimethyl Lys9. Lane 8: Trimethyl Lys9. Lane 9: Unmodified Lys27. Lane 10: Monomethyl Lys27. Lane 11: Dimethyl Lys27. Lane 12: Trimethyl Lys27.

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; it 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; they play a major role in regulating gene expression.

Histone H3K27 can be mono-, di- or trimethylated by different histone methyltransferases, such as EZH2 or NSD3. While histone methylation can be associated with transcriptional activation or repression, methylation of Lysine 27 of histone H3 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 trimethyl Lys27 antibody.

qPCR Primers

Active Motif's Human Positive Control Primer Set MYT1 (Cat. No. 71007) and Human Negative Control Primer Set 3 (Cat. No. 71023) have been validated for performing qPCR and endpoint PCR when this antibody has been used for ChIP on human samples. For ChIP experiments with this antibody on mouse samples, the Mouse Positive Control Primer Set Pax-2 (Cat. No. 71020) and Mouse Negative Control Primer Set 1 (Cat. No. 71011) have been validated for both qPCR and endpoint PCR.

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 H3K27me3 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;
  • WB = Western Blotting