Product nameAnti-KDM3A / JHDM2A antibody [14F8]
See all KDM3A / JHDM2A primary antibodies
DescriptionMouse monoclonal [14F8] to KDM3A / JHDM2A
Tested applicationsSuitable for: ChIP, Flow Cyt, IHC-P, WB, ELISA, IPmore details
Species reactivityReacts with: Mouse, Human
Recombinant fragment corresponding to Human KDM3A/ JHDM2A.
- Expressed KDM3A in 293T cells. Hela nuclear extracts for IP. This antibody gave a positive result in IHC in the following FFPE tissue: Human normal spleen.
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
Storage bufferConstituent: Tissue culture supernatant
PurityTissue culture supernatant
Our Abpromise guarantee covers the use of ab91252 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ChIP||Use at an assay dependent concentration. PubMed: 23386205|
|Flow Cyt||Use 1-2µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|IHC-P||Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
|WB||1/1000. Predicted molecular weight: 147 kDa.|
|ELISA||Use at an assay dependent concentration.|
|IP||Use at an assay dependent concentration.|
FunctionHistone demethylase that specifically demethylates 'Lys-9' of histone H3, thereby playing a central role in histone code. Preferentially demethylates mono- and dimethylated H3 'Lys-9' residue, with a preference for dimethylated residue, while it has weak or no activity on trimethylated H3 'Lys-9'. Demethylation of Lys residue generates formaldehyde and succinate. Involved in hormone-dependent transcriptional activation, by participating in recruitment to androgen-receptor target genes, resulting in H3 'Lys-9' demethylation and transcriptional activation. Involved in spermatogenesis by regulating expression of target genes such as PRM1 and TMP1 which are required for packaging and condensation of sperm chromatin. Involved in obesity resistance through regulation of metabolic genes such as PPARA and UCP1.
Sequence similaritiesBelongs to the JHDM2 histone demethylase family.
Contains 1 JmjC domain.
DomainThe JmjC domain and the C6-type zinc-finger are required for the demethylation activity.
Leu-Xaa-Xaa-Leu-Leu (LXXLL) motifs are known to mediate the association with nuclear receptors.
Cellular localizationCytoplasm. Nucleus. Nuclear in round spermatids. When spermatids start to elongate, localizes to the cytoplasm where it forms distinct foci which disappear in mature spermatozoa.
- Information by UniProt
- DKFZp686A24246 antibody
- DKFZp686P07111 antibody
- JHDM2A antibody
All lanes : Anti-KDM3A / JHDM2A antibody [14F8] (ab91252) at 1/1000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : KDM3A knockout HAP1 whole cell lysate
Lysates/proteins at 40 µg per lane.
Predicted band size: 147 kDa
Lanes 1 - 2: Merged signal (red and green). Green - ab91252 observed at 147 kDa. Red - loading control, ab52866, observed at 37 kDa.
ab91252 was shown to recognize in wild-type HAP1 cells as signal was lost at the expected MW in KDM3A knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and KDM3A knockout samples were subjected to SDS-PAGE. The membrane was blocked with 0. Ab91252 and ab52866 (Rabbit anti-alpha Tubulin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
IHC image of KDM3A / JHDM2A antibody staining in Human normal spleen formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab91252, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Cell extracts derived from 293T without (-) or with overexpressing Flag-KDM3A (+) were used for Western (left panel) and IP-Western (right panel) analysis of ab91252 (clone 14F8).
Regular IP buffer: 1mM EDTA; 150mM NaCl; 50mM TrisCl, pH 7.5; 1% Triton X-100; 2mM PMSF; Protease inhibitors ChIP C buffer: 1mM EDTA; 150mM NaCl; 50mM TrisCl, pH 7.5; 1% Triton X-100; 0.1% Sodium Deoxycholate; 2mM PMSF; Protease inhibitors
Overlay histogram showing HeLa cells stained with ab91252 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab91252, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
This product has been referenced in:
- Wang HY et al. Histone demethylase KDM3A is required for enhancer activation of hippo target genes in colorectal cancer. Nucleic Acids Res 47:2349-2364 (2019). Read more (PubMed: 30649550) »
- Brügger V et al. Delaying histone deacetylase response to injury accelerates conversion into repair Schwann cells and nerve regeneration. Nat Commun 8:14272 (2017). Read more (PubMed: 28139683) »