Overview

  • Product name
    Anti-KMT1E / SETDB1 antibody - ChIP Grade
    See all KMT1E / SETDB1 primary antibodies
  • Description
    Rabbit polyclonal to KMT1E / SETDB1 - ChIP Grade
  • Host species
    Rabbit
  • Tested applications
    Suitable for: ChIP, ICC/IF, WB, IHC-P, IPmore details
  • Species reactivity
    Reacts with: Mouse, Human
    Predicted to work with: Chimpanzee, Rhesus monkey, Gorilla, Orangutan
  • Immunogen

    Immunogen was a synthetic peptide, which represented a portion of human SET domain, bifurcated 1 encoded within exon 19 (LocusLink ID 9869).

  • Positive control
    • 293T cells.
  • General notes
    Centrifuge tube to remove product from lid.

Properties

Applications

Our Abpromise guarantee covers the use of ab12317 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ChIP Use at an assay dependent concentration. PubMed: 22110129
ICC/IF Use at an assay dependent concentration. PubMed: 21921037
WB 1/1000 - 1/10000. Predicted molecular weight: 155 kDa.
IHC-P Use at an assay dependent concentration.
IP Use a concentration of 1 - 4 mg/ml.

Target

  • Function
    Histone methyltransferase that specifically trimethylates 'Lys-9' of histone H3. H3 'Lys-9' trimethylation represents a specific tag for epigenetic transcriptional repression by recruiting HP1 (CBX1, CBX3 and/or CBX5) proteins to methylated histones. Mainly functions in euchromatin regions, thereby playing a central role in the silencing of euchromatic genes. H3 'Lys-9' trimethylation is coordinated with DNA methylation. Probably forms a complex with MBD1 and ATF7IP that represses transcription and couples DNA methylation and histone 'Lys-9' trimethylation. Its activity is dependent on MBD1 and is heritably maintained through DNA replication by being recruited by CAF-1. SETDB1 is targeted to histone H3 by TRIM28/TIF1B, a factor recruited by KRAB zinc-finger proteins.
  • Tissue specificity
    Widely expressed. High expression in testis.
  • Sequence similarities
    Belongs to the histone-lysine methyltransferase family. Suvar3-9 subfamily.
    Contains 1 MBD (methyl-CpG-binding) domain.
    Contains 1 post-SET domain.
    Contains 1 pre-SET domain.
    Contains 1 SET domain.
    Contains 2 Tudor domains.
  • Domain
    The pre-SET, SET and post-SET domains are all required for methyltransferase activity. The 347-amino-acid insertion in the SET domain has no effect on the catalytic activity.
    Isoform 2 lacks all domains required for histone methyltransferase activity.
  • Cellular localization
    Nucleus. Chromosome. Associated with non-pericentromeric regions of chromatin. Excluded from nucleoli and islands of condensed chromatin.
  • Information by UniProt
  • Database links
  • Alternative names
    • AU022152 antibody
    • EC 2.1.1.43 antibody
    • ERG-associated protein with SET domain antibody
    • ESET antibody
    • H3 K9 HMTase 4 antibody
    • H3-K9-HMTase 4 antibody
    • H3-K9-HMTase4 antibody
    • Histone H3 K9 methyltransferase 4 antibody
    • Histone H3-K9 methyltransferase 4 antibody
    • Histone-lysine N-methyltransferase SETDB1 antibody
    • KG1T antibody
    • KIAA0067 antibody
    • KMT1E antibody
    • Lysine N-methyltransferase 1E antibody
    • MGC90670 antibody
    • mKIAA0067 antibody
    • SET domain bifurcated 1 antibody
    • Set domain protein, bifurcated, 1 antibody
    • SETB1_HUMAN antibody
    • Setdb1 antibody
    see all

Images

  • Samples: Whole cell lysate (10 mcg for 3 and 4; 50 mcg for 1 and 2) from 293T cells that were mock transfected (1 and 2) or transfected with a SETDB1 expression construct (3 and 4).
  • ab12317 (4ug/ml) staining KMT1E in human testis using an automated system (DAKO Autostainer Plus). Using this protocol there is strong staining of KMT1E in nuclear compartments of the germinal epithelium.
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
  • Whole cell lysate (2 mg) from 293T cells that were mock transfected (1 and 2) or transfected with a SETDB1 expression construct (3 and 4). Rabbit anti-SETDB1 antibodies; ab12317 (2 and 4) and a competitor's antiserum (1 and 3) were used at 1 mcg/mg lysate for IP. ab12317 was used at 0.2 mcg/ml for Western Blot. Detection: Chemiluminescence with exposure times of 1 to 5 minutes.

References

This product has been referenced in:
  • Alarcon V  et al. The enzymes LSD1 and Set1A cooperate with the viral protein HBx to establish an active hepatitis B viral chromatin state. Sci Rep 6:25901 (2016). Read more (PubMed: 27174370) »
  • Hatanaka Y  et al. Histone chaperone CAF-1 mediates repressive histone modifications to protect preimplantation mouse embryos from endogenous retrotransposons. Proc Natl Acad Sci U S A 112:14641-6 (2015). ICC/IF ; Mouse . Read more (PubMed: 26546670) »
See all 17 Publications for this product

Customer reviews and Q&As

1-7 of 7 Abreviews or Q&A

Application
Immunohistochemistry (Frozen sections)
Sample
Human Tissue sections (brain)
Permeabilization
Yes - 0.5% Triton
Specification
brain
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 4°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Sep 01 2015

Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 37°C
Sample
Mouse Tissue sections (Ovary section)
Specification
Ovary section
Permeabilization
Yes - Trition
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Jan 30 2015

Answer

Thank you for taking time to answer the additional questions. I am sorry to hear this antibody is not providing satisfactory results.
The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality. I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful.
Having reviewed the protocol details, I believe this product should have given satisfactory results.
I apologize for the inconvenience and am pleased to offer you a free of charge replacement, credit note, or refund in compensation. Can you please confirm that your order number?
Thank you for your cooperation. I look forward to hearing from you with details of how you would like to proceed.

Read More

Answer

Thanks for your enquiry. There are two western blots on the datasheet for ab12317. I believe you are referring to the bottom image. I think there is a mistake in the annotation for this blot. 'Mock transfected' should refer to lanes 3 and 4 of this blot, not lanes 1 and 2. I am sorry for the confusion here. I hope this information helps to explain things. Please feel free to contact me again with any further questions.

Read More

Answer

Thank you for your enquiry. The predicted molecular weight of SETDB1 is 155kDa, based on the amino acid composition of the protein. It should be noted however that this protein is known to be phophorylated, as indicated on the Swissprot data page entry for SETDB1 under the 'Sequence annotation (Features)' section: http://www.uniprot.org/uniprot/Q15047 Phosphorylation of a protein will slow its migration through a gel, making it appear much heavier than normal. This may be what you are seeing. Alternatively, sometimes proteins run a little slower than expected based on their amino acid composition, even without post translational modifications. This may be what is happening here. Similar results are seen with ab12317, another antibody to SETDB1. ab12317 detects a band of 170kDa via western blotting. I hope this is helpful. Feel free to send me an image of your blot and I'll be happy to look at it.  Please contact me again if you have any further questions.

Read More
Abcam has not validated the combination of species/application used in this Abreview.
Application
ChIP on chip
Sample
Mouse Cell lysate - nuclear (mouse ES cells)
Specification
mouse ES cells
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 15 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% Formaldehyde
Detection step
Microarray
Positive control
Competitor Antibody

Dr. Steve Bilodeau

Verified customer

Submitted Oct 21 2009

Answer

The precise sequence used to generate ab12317 is proprietary. The epitope recognized by ab12317 is found between residues 1125 and 1175 of SwissProt entry Q15047 (http://us.expasy.org/cgi-bin/niceprot.pl?Q15047).

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