Overview

  • Product name
    Anti-Dnmt3a antibody [EPR18455]
    See all Dnmt3a primary antibodies
  • Description
    Rabbit monoclonal [EPR18455] to Dnmt3a
  • Host species
    Rabbit
  • Tested applications
    Suitable for: Flow Cyt, WB, ICC/IF, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human Dnmt3a aa 600-700. The exact sequence is proprietary.
    Database link: Q9Y6K1

  • Positive control
    • WB: HeLa, HEK-293 and C6 cell lysates; Rat brain and heart lysates. ICC/IF: HeLa cells. IHC-P: Human placenta, mouse testis, rat spleen tissue
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.01% Sodium azide
    Constituents: PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity
    Protein A purified
  • Clonality
    Monoclonal
  • Clone number
    EPR18455
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab188470 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
Flow Cyt Use at an assay dependent concentration.
WB 1/2000. Detects a band of approximately 130 kDa (predicted molecular weight: 102 kDa).
ICC/IF 1/1000.
IHC-P 1/2000.

Target

  • Function
    Required for genome wide de novo methylation and is essential for the establishment of DNA methylation patterns during development. DNA methylation is coordinated with methylation of histones. It modifies DNA in a non-processive manner and also methylates non-CpG sites. May preferentially methylate DNA linker between 2 nucleosomal cores and is inhibited by histone H1. Plays a role in paternal and maternal imprinting. Required for methylation of most imprinted loci in germ cells. Acts as a transcriptional corepressor for ZNF238. Can actively repress transcription through the recruitment of HDAC activity.
  • Tissue specificity
    Highly expressed in fetal tissues, skeletal muscle, heart, peripheral blood mononuclear cells, kidney, and at lower levels in placenta, brain, liver, colon, spleen, small intestine and lung.
  • Sequence similarities
    Belongs to the C5-methyltransferase family.
    Contains 1 ADD domain.
    Contains 1 GATA-type zinc finger.
    Contains 1 PHD-type zinc finger.
    Contains 1 PWWP domain.
  • Domain
    The PWWP domain is essential for targeting to pericentric heterochromatin.
  • Post-translational
    modifications
    Sumoylated; sumoylation disrupts the ability to interact with histone deacetylases (HDAC1 and HDAC2) and repress transcription.
  • Cellular localization
    Nucleus. Cytoplasm. Accumulates in the major satellite repeats at pericentric heterochromatin.
  • Information by UniProt
  • Database links
  • Alternative names
    • DNA (cytosine 5) methyltransferase 3 alpha antibody
    • DNA (cytosine 5) methyltransferase 3A antibody
    • DNA (cytosine-5)-methyltransferase 3A antibody
    • DNA cytosine methyltransferase 3A2 antibody
    • DNA methyltransferase 3 alpha antibody
    • DNA methyltransferase 3a antibody
    • DNA methyltransferase HsaIIIA antibody
    • DNA MTase HsaIIIA antibody
    • DNM3A_HUMAN antibody
    • DNMT 3a antibody
    • DNMT antibody
    • Dnmt3a antibody
    • DNMT3A2 antibody
    • M.HsaIIIA antibody
    • MCMT antibody
    • OTTHUMP00000201149 antibody
    • TBRS antibody
    see all

Images

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: Dnmt3a knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (20 µg)
    Lane 4: Mouse brain tissue lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab188470 observed at 125 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab188470 was shown to recognize Dnmt3a when Dnmt3a knockout samples were used, along with additional cross-reactive bands. Wild-type and Dnmt3a knockout samples were subjected to SDS-PAGE. ab188470 and ab8245 (loading control to GAPDH) were diluted to 1/5000 and 1/10000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human placenta tissue sections labeling Dnmt3a with purified ab188470 at 1/2000 (0.409 µg/ml). Antigen retrieval was heat mediated using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counterstain.

  • Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Dnmt3a with purified ab188470 at 1/80 dilution(10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488)(1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

  • All lanes : Anti-Dnmt3a antibody [EPR18455] (ab188470) at 1/5000 dilution

    Lane 1 : HeLa whole cell lysate
    Lane 2 : Mouse skin tissue lysate
    Lane 3 : Rat skin tissue lysate
    Lane 4 : Mouse brain tissue lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size: 102 kDa
    Observed band size: 125 kDa
    why is the actual band size different from the predicted?



    Lanes 1 - 4: Merged signal (red and green). Green - ab188470 observed at 125 kDa. Red - loading control, ab8245, observed at 37 kDa.

     

    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab188470 and ab8245 (loading control) overnight at 4°C. Antibody binding was detected using Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at a 1:10000 dilution for 1hr at room temperature and then imaged.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat spleen tissue sections labeling Dnmt3 with purified ab188470 at 1/2000 (0.409 µg/ml). Antigen retrieval was heat mediated using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counterstain.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse testis tissue sections labeling Dnmt3a with purified ab188470 at 1/2000 (0.409 µg/ml). Antigen retrieval was heat mediated using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counterstain.

  • Anti-Dnmt3a antibody [EPR18455] (ab188470) at 1/10000 dilution + HeLa (Human epithelial cells from cervix adenocarcinoma) cell lysate at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

    Predicted band size: 102 kDa
    Observed band size: 130 kDa why is the actual band size different from the predicted?


    Exposure time: 5 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed MW is consistent with what has been described in the literature (J Biol Chem. 2002. 277, 38746-38754. PMID: 2138111).

  • Anti-Dnmt3a antibody [EPR18455] (ab188470) at 1/10000 dilution + HEK-293 (Human epithelial cells from embryonic kidney) cell lysate at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

    Predicted band size: 102 kDa
    Observed band size: 130 kDa why is the actual band size different from the predicted?


    Exposure time: 15 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed MW is consistent with what has been described in the literature (J Biol Chem. 2002. 277, 38746-38754. PMID: 2138111).

  • All lanes : Anti-Dnmt3a antibody [EPR18455] (ab188470) at 1/2000 dilution

    Lane 1 : Rat brain lysate
    Lane 2 : Rat heart lysate
    Lane 3 : C6 (Rat glial tumor cells) cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

    Predicted band size: 102 kDa
    Observed band size: 130 kDa why is the actual band size different from the predicted?


    Exposure time: 30 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling Dnmt3a with ab188470 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear and weakly cytoplasmic staining on HeLa cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab188470 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
    -ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

References

This product has been referenced in:
  • Zhang S  et al. Up-regulation of microRNA-203 in influenza A virus infection inhibits viral replication by targeting DR1. Sci Rep 8:6797 (2018). WB . Read more (PubMed: 29717211) »
  • Zhang M  et al. Blockade of receptors of advanced glycation end products ameliorates diabetic osteogenesis of adipose-derived stem cells through DNA methylation and Wnt signalling pathway. Cell Prolif 51:e12471 (2018). Read more (PubMed: 30014569) »
See all 4 Publications for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Astatotilapia burtoni Tissue sections (thalamus/hypothalamus)
Antigen retrieval step
None
Permeabilization
Yes - 0.2% TritonX
Specification
thalamus/hypothalamus
Blocking step
10% normal serum 1% BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C
Fixative
Paraformaldehyde

Sebastian Alvarado

Verified customer

Submitted Dec 18 2015

Application
Western blot
Sample
Astatotilapia burtoni Cell lysate - whole cell (Brain and gonads)
Gel Running Conditions
Reduced Denaturing (5% acrylamide gel)
Loading amount
25 µg
Specification
Brain and gonads
Blocking step
Milk as blocking agent for 15 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Sebastian Alvarado

Verified customer

Submitted Aug 07 2015

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