Overview

  • Product name

    Anti-LINE-1 ORF1p antibody [EPR22227-54]
    See all LINE-1 ORF1p primary antibodies
  • Description

    Rabbit monoclonal [EPR22227-54] 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: K562, MCF7 and NCCIT whole cell lysates. IP: NCCIT whole cell lysate. IHC-P: Human lung cancer, pancreatic cancer and colon cancer tissue. ICC/IF: NCCIT and MCF7 cells. Flow cyt: NCCIT cells.
  • 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

    EPR22227-54
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab230966 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. Detects a band of approximately 33, 42 kDa (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 MCF7 (human breast adenocarcinoma cell line) cells labeling LINE-1 ORF1p with ab230966 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). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 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 NCCIT (human pluripotent embryonic carcinoma cell line) cells labeling LINE-1 ORF1p with ab230966 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: 15028673; 17562864). The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 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 tissue (Panel A) and adjacent human normal colon tissue (Panel B) labeling LINE-1 ORF1p with ab230966 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Cytoplasmic staining in colon cancer (Panel A) but negative in adjacent normal tissue (Panel B). (PMID: 24607009) is observed. Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

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

  • Immunohistochemical analysis of paraffin-embedded human pancreatic cancer tissue (Panel A) and adjacent human normal pancreatic tissue (Panel B) labeling LINE-1 ORF1p with ab230966 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Cytoplasmic staining in pancreatic cancer (Panel A) but negative in adjacent normal tissue (Panel B). (PMID: 24607009) is observed. Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

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

  • Immunohistochemical analysis of paraffin-embedded human lung cancer tissue (Panel A) and adjacent human normal lung tissue (Panel B) labeling LINE-1 ORF1p with ab230966 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Cytoplasmic staining in lung cancer (Panel A) but negative in adjacent normal tissue (Panel B) (PMID: 24607009) is observed. Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

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

  • LINE-1 ORF1p was immunoprecipitated from 0.35 mg of NCCIT (human pluripotent embryonic carcinoma cell line) whole cell lysate with ab230966 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab230966 at 1/1000 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: ab230966 IP in NCCIT whole cell lysate. 
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab230966 in NCCIT whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.
    Exposure time: 1 second.

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

  • Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HCT 116 (human colorectal carcinoma cell line)(Left panel) and NCCIT (human pluripotent embryonic carcinoma cell line) (Right panel) cells labeling LINE-1 ORF1p with ab230966 at 1/500 dilution (red) compared with Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077), at 1/2000 dilution was used as the secondary antibody.

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

  • All lanes : Anti-LINE-1 ORF1p antibody [EPR22227-54] (ab230966) at 1/1000 dilution

    Lane 1 : HCT 116 (human colorectal carcinoma cell line) whole cell lysate
    Lane 2 : K562 (human chronic myelogenous leukemia cell line from bone marrow ) whole cell lysate
    Lane 3 : MCF7 (human breast adenocarcinoma cell line) whole cell lysate
    Lane 4 : NCCIT (human pluripotent embryonic carcinoma cell line) 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?


    Exposure time: 1 second


    Blocking and dilution buffer: 5% NFDM/TBST.

    Exposure times.

    Lanes 1 and 2: 37 seconds.

    Lanes 3 and 4: 15 seconds.

    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).

References

ab230966 has not yet been referenced specifically in any publications.

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