Overview

  • Product name

    Anti-KDM1/LSD1 antibody [EPR6825] - Nuclear Marker
    See all KDM1/LSD1 primary antibodies
  • Description

    Rabbit Monoclonal [EPR6825] to KDM1/LSD1 - Nuclear Marker
  • Host species

    Rabbit
  • Tested applications

    Suitable for: Flow Cyt, WB, IP, IHC-P, ICC/IF, ChIPmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human KDM1/ LSD1 aa 50-150. The exact sequence is proprietary.
    Database link: O60341
    (Peptide available as ab166919)

  • Positive control

    • 293T, HeLa, Jurkat, PC3, C6, RAW264.7, PC12, and NIH 3T3 cell lysates; Human testis tissue; HeLa cells. ICC/IF: HAP1-KDM1A cells. ChIP: HCT 116( Human colorectal carcinoma epithelial cell)
  • 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.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    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. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR6825
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab129195 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 1/20.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

WB 1/10000 - 1/50000. Detects a band of approximately 110 kDa (predicted molecular weight: 92 kDa).Can be blocked with KDM1 / LSD1 peptide (ab166919).
IP 1/10 - 1/100.
IHC-P 1/50 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ICC/IF Use a concentration of 1 µg/ml.
ChIP Use 5 µg for 25 µg of chromatin.

Target

  • Function

    Histone demethylase that demethylates both 'Lys-4' (H3K4me) and 'Lys-9' (H3K9me) of histone H3, thereby acting as a coactivator or a corepressor, depending on the context. Acts by oxidizing the substrate by FAD to generate the corresponding imine that is subsequently hydrolyzed. Acts as a corepressor by mediating demethylation of H3K4me, a specific tag for epigenetic transcriptional activation. Demethylates both mono- (H3K4me1) and di-methylated (H3K4me2) H3K4me. May play a role in the repression of neuronal genes. Alone, it is unable to demethylate H3K4me on nucleosomes and requires the presence of RCOR1/CoREST to achieve such activity. Also acts as a coactivator of androgen receptor (ANDR)-dependent transcription, by being recruited to ANDR target genes and mediating demethylation of H3K9me, a specific tag for epigenetic transcriptional repression. The presence of PRKCB in ANDR-containing complexes, which mediates phosphorylation of 'Thr-6' of histone H3 (H3T6ph), a specific tag that prevents demethylation H3K4me, prevents H3K4me demethylase activity of KDM1A. Demethylates di-methylated 'Lys-370' of p53/TP53 which prevents interaction of p53/TP53 with TP53BP1 and represses p53/TP53-mediated transcriptional activation. Demethylates and stabilizes the DNA methylase DNMT1. Required for gastrulation during embryogenesis.
  • Tissue specificity

    Ubiquitously expressed.
  • Sequence similarities

    Belongs to the flavin monoamine oxidase family.
    Contains 1 SWIRM domain.
  • Domain

    The SWIRM domain may act as an anchor site for a histone tail.
  • Cellular localization

    Nucleus.
  • Information by UniProt
  • Database links

  • Alternative names

    • Amine oxidase (flavin containing) domain 2 antibody
    • Amine oxidase, flavin containing, 2 antibody
    • AOF2 antibody
    • BHC110 antibody
    • BRAF35 HDAC complex protein BHC110 antibody
    • BRAF35-HDAC complex protein BHC110 antibody
    • BRAF35/HDAC complex, 110-kD subunit antibody
    • CPRF antibody
    • EC1 antibody
    • FAD binding protein BRAF35 HDAC complex, 110 kDa subunit antibody
    • Flavin-containing amine oxidase domain-containing protein 2 antibody
    • KDM 1 antibody
    • KDM1 antibody
    • Kdm1a antibody
    • KDM1A_HUMAN antibody
    • KIAA0601 antibody
    • LSD 1 antibody
    • LSD1 antibody
    • Lysine (K) specific demethylase 1 antibody
    • Lysine (K) specific demethylase 1A antibody
    • Lysine demethylase 1A antibody
    • Lysine specific histone demethylase 1 antibody
    • Lysine specific histone demethylase 1A antibody
    • Lysine-specific demethylase 1 antibody
    • Lysine-specific demethylase 1A antibody
    • Lysine-specific histone demethylase 1A antibody
    see all

Images

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: KMD1 / LSD1 knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (20 µg)
    Lane 4: Jurkat cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab129195 observed at 110 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab129195 was shown to specifically react with KMD1 / LSD1 in wild-type HAP1 cells. No band was observed when KMD1 / LSD1 knockout samples were used. Wild-type and KMD1 / LSD1 knockout samples were subjected to SDS-PAGE. ab129195 and ab8245 (loading control to GAPDH) were both diluted 1/10,000 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 1/10,000 dilution for 1 hour at room temperature before imaging.

  • Chromatin was prepared from HCT 116 cells according to the Abcam X-ChIP protocol. Cells were fixed with 1% formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 5µg of ab129195 (red), and 20µl of protein A/G sepharose beads slurry (10µl of sepharose A beads + 10µl of sepharose G beads). 5μg of rabbit normal IgG was added to the beads control (grey). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

  • ab129195 staining KDM1A/LSD1 in wild-type HAP1 cells (top panel) and KDM1A knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab129195 at 1μg/ml concentration and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

  • Immunohistochemical staining of paraffin embedded rat kidney with purified ab129195 at a working dilution of 1/50. The secondary antibody used is ab97051, a goat anti-rabbit IgG (H&L) at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
  • Immunofluorescence staining of HeLa cells with purified ab129195 at a working dilution of 1/100, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab129195 was used at a dilution of 1/500 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

  • Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling KDM1/LSD1 with purified ab129195 at 1/20 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.

  • ab129195 (purified) at 1/20 immunoprecipitating KDM1/LSD1 in 10 μg Jurkat cell lysate (Lanes 1 and 2, observed at 110 kDa). Lane 3 - Rabbit monoclonal IgG (ab172730). For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution. Blocking buffer and concentration: 5% NFDM/TBST Dilution buffer and concentration: 5% NFDM/TBST

  • All lanes : Anti-KDM1/LSD1 antibody [EPR6825] - Nuclear Marker (ab129195) at 1/10000 dilution (purified)

    Lane 1 : C6 whole cell lysate
    Lane 2 : Raw 264.7 whole cell lysate
    Lane 3 : PC-12 whole cell lysate
    Lane 4 : NIH/3T3 whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (HRP goat anti-rabbit IgG (H+L))

    Predicted band size: 92 kDa
    Observed band size: 110 kDa
    why is the actual band size different from the predicted?



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Immunohistochemical staining of paraffin embedded mouse colon with purified ab129195 at a working dilution of 1/50. The secondary antibody used is ab97051, a goat anti-rabbit IgG (H&L) at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
  • All lanes : Anti-KDM1/LSD1 antibody [EPR6825] - Nuclear Marker (ab129195) at 1/10000 dilution (purified)

    Lane 1 : HEK293 whole cell lysate
    Lane 2 : HeLa whole cell lysate
    Lane 3 : Jurkat whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (HRP goat anti-rabbit IgG (H+L))

    Predicted band size: 92 kDa
    Observed band size: 110 kDa why is the actual band size different from the predicted?



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Immunohistochemical staining of paraffin embedded human stomach carcinoma with purified ab129195 at a working dilution of 1/50. The secondary antibody used is ab97051, a goat anti-rabbit IgG (H&L) at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
  • Unpurified ab129195 staining KDM1/LSD1 in human paraffin-embedded A549 lung cancer cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed using the HOPE technique and permeabilized with 0.05% Tween. Samples were incubated with primary antibody (1/100) for 45 minutes at 25°C. An Alexa Fluor®488-conjugated Donkey anti-mouse IgG polyclonal (1/200) was used as the secondary antibody.

    See Abreview

  • All lanes : Anti-KDM1/LSD1 antibody [EPR6825] - Nuclear Marker (ab129195) at 1/10000 dilution (unpurified)

    Lane 1 : 293T cell lysate
    Lane 2 : HeLa cell lysate
    Lane 3 : Jurkat cell lysate
    Lane 4 : PC3 cell lysate
    Lane 5 : C6 cell lysate
    Lane 6 : RAW264.7 cell lysate
    Lane 7 : PC12 cell lysate
    Lane 8 : NIH 3T3 cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat anti-Rabbit HRP at 1/2000 dilution

    Predicted band size: 92 kDa

  • Unpurified ab129195, at 1/100, staining KDM1 / LSD1 in paraffin embedded Human testis tissue by Immunohistochemistry.

    Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

References

This product has been referenced in:

  • Shen L  et al. TLR4/NF-?B axis signaling pathway-dependent up-regulation of miR-625-5p contributes to human intervertebral disc degeneration by targeting COL1A1. Am J Transl Res 11:1374-1388 (2019). Read more (PubMed: 30972168) »
  • Kilinc S  et al. Lysine-specific demethylase-1 (LSD1) is compartmentalized at nuclear chromocenters in early post-mitotic cells of the olfactory sensory neuronal lineage. Mol Cell Neurosci 74:58-70 (2016). IF, IHC, WB, IP ; Mouse . Read more (PubMed: 26947098) »
See all 4 Publications for this product

Customer reviews and Q&As

1-6 of 6 Abreviews or Q&A

Application
Western blot
Sample
Mouse Cell lysate - whole cell (mouse embryonic stem cells)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
15 µg
Specification
mouse embryonic stem cells
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C

Abcam user community

Verified customer

Submitted Jan 25 2016

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Permeabilization
Yes - 0.5% Triton-X100 in PBS
Specification
HeLa
Fixative
Paraformaldehyde

Dr. Kirk Mcmanus

Verified customer

Submitted Feb 25 2015

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (Paraffin-embedded A549 lung cancer cells)
Permeabilization
Yes - 0.05 % TWEEN in wash buffer
Specification
Paraffin-embedded A549 lung cancer cells
Fixative
HOPE

Abcam user community

Verified customer

Submitted Apr 30 2014

Answer

Thank you for your enquiry regarding ab129195 and for taking the time to provide some useful details of the experiments. I am very sorry to hear that you are having problems with this antibody.
After reading through the detailed protocol you kindly forwarded to Abcam, I would like to make the following comment/suggestion:
This antibody ab129195: Rabbit monoclonal [EPR6825] to KDM1 / LSD1 so the compatible secondary antibody should be anti-rabbit and not anti-mouse. I would advise you to check the compatibility since this may have caused the problem.
If you need any further assistance in the future, please do not hesitate to contact me.

Read More

Answer


As for the blocking peptides:

For ab129195: The blocking peptide is now available as ab166919.
https://www.abcam.com/index.html?datasheet=166919

For ab62582: This will be available soon and I will let you know when I have a catalog number and price.

Read More
Application
Western blot
Sample
Human Cell lysate - other (HeLa)
Gel Running Conditions
Reduced Denaturing
Loading amount
20 µg
Specification
HeLa
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 16°C

Abcam user community

Verified customer

Submitted Jan 10 2013

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