Overview

  • Product name

    Anti-LINE-1 ORF1p antibody [EPR22227-6]
    See all LINE-1 ORF1p primary antibodies
  • Description

    Rabbit monoclonal [EPR22227-6] to LINE-1 ORF1p
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Human
  • Immunogen

    Recombinant fragment within Human LINE-1 ORF1p aa 150 to the C-terminus. The exact sequence is proprietary.
    Database link: Q9UN81

  • Positive control

    • WB: NCCIT, MCF7 and K562 whole cell lysate. IHC-P: Human pancreatic cancer, colon cancer and lung cancer tissue. ICC/IF: NCCIT and MCF7 cells. Flow cyt: NCCIT cells. IP: NCCIT whole cell lysate.
  • 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

    EPR22227-6
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab245249 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
WB 1/1000. Predicted molecular weight: 40 kDa.
IHC-P 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF 1/100.
Flow Cyt 1/500.
IP 1/30.

Target

  • Relevance

    Domain: The coiled coil domain mediates homotrimerization. The RRM and the CTD domain are both required for proper RNA-binding activity. Function: Nucleic acid-binding protein which is essential for retrotransposition of LINE-1 elements in the genome. Functions as a nucleic acid chaperone binding its own transcript and therefore preferentially mobilizing the transcript from which they are encoded. Long interspersed element-1/LINE-1/L1 retrotransposons are present in more than 500'000 full (6 kb) or truncated copies in the human genome. Most of them are inactive but 80 to 100 of those elements could be transcribed, translated and active in any individual. An active LINE-1 encodes for 2 proteins translated from a single RNA containing 2 non-overlapping ORFs, ORF1 and ORF2. ORF1p is described in this entry as a representative of all ORF1p potentially expressed by active elements. ORF2p is described in the related entry AC O00370. PTM: Polyubiquitinated, probably by UBR2, which induces its degradation. Similarity: Belongs to the transposase 22 family.
  • Database links

  • Alternative names

    • L1ORF1p antibody
    • L1RE1 antibody
    • LINE retrotransposable element 1 antibody
    • LINE-1 retrotransposable element ORF1 protein antibody
    • LINE1 retrotransposable element 1 antibody
    • LRE1 antibody
    see all

Images

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NCCIT (human pluripotent embryonic carcinoma epithelial cell) cells labeling LINE-1 ORF1p with ab245249 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic with punctate foci and nucleolar staining in NCCIT cells (PMID: 17562864; 15028673). The nuclear counter stain is DAPI (blue).
    Counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at a 1/200 dilution (red).
    Secondary antibody only control:  Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized MCF7 (human breast adenocarcinoma epithelial cell) cells labeling LINE-1 ORF1p with ab245249 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic with punctate foci and nucleolar staining in MCF7 cells (PMID: 15028673; 17562864). The nuclear counter stain is DAPI (blue).
    Counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at a 1/200 dilution (red).
    Secondary antibody only control:  Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution..

  • Immunohistochemical analysis of paraffin-embedded human colon cancer (Panel A) and adjacent normal tissue (Panel B) labeling LINE-1 ORF1p with ab245249 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining in colon cancer (Panel A) but negative in adjacent normal tissue (Panel B). (PMID:24607009). Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).
    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Immunohistochemical analysis of paraffin-embedded human pancreatic cancer (Panel A) and adjacent normal tissue (Panel B) labeling LINE-1 ORF1p with ab245249 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining in pancreatic cancer (Panel A) but negative in adjacent normal tissue (Panel B). (PMID:24607009). Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Immunohistochemical analysis of paraffin-embedded human lung cancer (Panel A) and adjacent normal tissue (Panel B) labeling LINE-1 ORF1p with ab245249 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining in lung cancer (Panel A) but negative in adjacent normal tissue (Panel B). (PMID:24607009). Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • LINE-1 ORF1p was immunoprecipitated from 0.35 mg NCCIT (human pluripotent embryonic carcinoma epithelial cell) whole cell lysate with ab245249 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab245249 at 1/2000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

    Lane 1: NCCIT whole cell lysate 10 µg (Input).
    Lane 2: ab245249 IP in NCCIT whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab245249 in NCCIT whole cell lysate.

    The lower 33-kDa band is likely to be the truncated form (PMID: 29505568).

    Blocking/Dilution buffer: 5% NFDM/TBST.
    Exposure time: 1 second.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HCT 116 (human colorectal carcinoma epithelial cell, Left panel) / NCCIT (human pluripotent embryonic carcinoma epithelial cell, Right panel) cell lines labeling LINE-1 ORF1p with ab245249 at 1/500 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

    The expression level on HCT 116 (low) and NCCIT (high) is consistent with literature (PMID: 27016617).

  • All lanes : Anti-LINE-1 ORF1p antibody [EPR22227-6] (ab245249) at 1/1000 dilution

    Lane 1 : NCCIT (human pluripotent embryonic carcinoma epithelial cell) whole cell lysate
    Lane 2 : MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate
    Lane 3 : K562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate
    Lane 4 : HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 40 kDa
    Observed band size: 33,42 kDa why is the actual band size different from the predicted?



    The molecular weight observed is consistent with what has been described in the literature (PMID: 29505568).

    The lower 33-kDa band is likely to be a truncated form (PMID: 29505568).

    Control cell line: HCT-116 (low-expression, PMID: 27016617).

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure times: Lane 1: 5 seconds; Lane 2, 3 and 4: 37 seconds.

References

ab245249 has not yet been referenced specifically in any publications.

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