Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Granulin antibody [EPR15864] (ab208777)

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Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR15864] to Granulin
  • Suitable for: WB, IP, IHC-P
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-Granulin antibody [EPR15864]
    See all Granulin primary antibodies

  • Description

    Rabbit monoclonal [EPR15864] to Granulin

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, IP, IHC-Pmore details

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant fragment within Human Granulin aa 100-250. The exact sequence is proprietary.
    Database link: P28799

  • Positive control

    • WB: HepG2, HAP1, A431 and 293T whole cell lysate; Human fetal kidney lysate; Human GRN full-length recombinant protein with GST-tag at N-terminal. IHC-P: Human liver, placenta, breast cancer and gastric cancer tissues. IP: HeLa whole cell lysate.

  • General notes

     

     

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

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

Properties

Applications

Our Abpromise guarantee covers the use of ab208777 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 1/2000. Detects a band of approximately 74 kDa (predicted molecular weight: 64 kDa).
IP 1/70.
IHC-P 1/1000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Target

  • Function

    Granulins have possible cytokine-like activity. They may play a role in inflammation, wound repair, and tissue remodeling.
    Granulin-4 promotes proliferation of the epithelial cell line A431 in culture while granulin-3 acts as an antagonist to granulin-4, inhibiting the growth.

  • Tissue specificity

    In myelogenous leukemic cell lines of promonocytic, promyelocytic, and proerythroid lineage, in fibroblasts, and very strongly in epithelial cell lines. Present in inflammatory cells and bone marrow. Highest levels in kidney.

  • Involvement in disease

    Defects in GRN are the cause of ubiquitin-positive frontotemporal dementia (UP-FTD) [MIM:607485]; also known as tau-negative frontotemporal dementia linked to chromosome 17. Frontotemporal dementia (FTD) is the second most common cause of dementia in people under the age of 65 years. It is an autosomal dominant neurodegenerative disease.

  • Sequence similarities

    Belongs to the granulin family.

  • Post-translational
    modifications

    Granulins are disulfide bridged.

  • Cellular localization

    Secreted.
  • Information by UniProt

  • Database links

    • Alternative names

      • Acrogranin antibody
      • CLN11 antibody
      • GEP antibody
      • GP88 antibody
      • Granulin A antibody
      • Granulin B antibody
      • Granulin C antibody
      • Granulin D antibody
      • Granulin E antibody
      • Granulin epithelin antibody
      • Granulin F antibody
      • Granulin G antibody
      • Granulin-7 antibody
      • Granulins antibody
      • GRN antibody
      • GRN_HUMAN antibody
      • PC cell derived growth factor antibody
      • PCDGF antibody
      • PEPI antibody
      • PGRN antibody
      • Proepithelin antibody
      • Progranulin antibody
      see all

    Images

    • Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      All lanes : Anti-Granulin antibody [EPR15864] (ab208777) at 1/1000 dilution

      Lane 1 : Wild-type HEK293T cell lysate
      Lane 2 : GRN knockout HEK293T cell lysate
      Lane 3 : HepG2 cell lysate
      Lane 4 : HAP1 cell 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

      Predicted band size: 64 kDa
      Observed band size: 74 kDa
      why is the actual band size different from the predicted?



      Lanes 1-4: Merged signal (red and green). Green - ab208777 observed at 74 kDa. Red - loading control ab8245 observed at 36 kDa.

       ab208777 Anti-Granulin antibody [EPR15864] was shown to specifically react with Granulin in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266738 (knockout cell lysate ab257235) was used. Wild-type and Granulin knockout samples were subjected to SDS-PAGE. ab208777 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2.5 hours at 1 in 1000 dilution and 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)

      Immunohistochemical analysis of paraffin-embedded Human placenta tissue labeling Granulin with ab208777 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on trophoblastic cells of Human placenta is observed. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunoprecipitation - Anti-Granulin antibody [EPR15864] (ab208777)
      Immunoprecipitation - Anti-Granulin antibody [EPR15864] (ab208777)

      Granulin was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab208777 at 1/70 dilution.

      Western blot was performed from the immunoprecipitate using ab208777 at 1/1000 dilution.

      VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

      Lane 1: HeLa whole cell lysate, 10µg (Input).

      Lane 2: ab208777 IP in HeLa whole cell lysate.

      Lane 3: Rabbit IgG,monoclonal -Isotype Control (ab172730) instead of ab208777 in HeLa whole cell lysate.

      Blocking and dilution buffer and concentration: 5% NFDM/TBST.

      Exposure time: 30 seconds.

    • Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      All lanes : Anti-Granulin antibody [EPR15864] (ab208777) at 1/2000 dilution

      Lane 1 : Wild-type HAP1 whole cell lysate
      Lane 2 : GRN (Granulin) knockout HAP1 whole cell lysate
      Lane 3 : HepG2 whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Predicted band size: 64 kDa



      Lanes 1 - 3: Merged signal (red and green). Green - ab208777 observed at 64 kDa. Red - loading control, ab8245, observed at 37 kDa.

      ab208777 was shown to specifically react with Granulin in wild-type HAP1 cells as signal was lost in GRN (Granulin) knockout cells. Wild-type and GRN (Granulin) knockout samples were subjected to SDS-PAGE. Ab208777 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/2000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

    • Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Lane 1 : Anti-Granulin antibody [EPR15864] (ab208777) at 1/2000 dilution
      Lane 2 : Anti-Granulin antibody [EPR15864] (ab208777) at 1/10000 dilution

      Lane 1 : A431 (Human epidermoid carcinoma cell line) whole cell lysate
      Lane 2 : 293T (Human epithelial cell line from embryonic kidney) whole cell lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

      Predicted band size: 64 kDa
      Observed band size: 74 kDa why is the actual band size different from the predicted?


      Exposure time: 1 minute


      Blocking/Dilution buffer: 5% NFDM/TBST.

    • Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Anti-Granulin antibody [EPR15864] (ab208777) at 1/2000 dilution + Human fetal kidney lysate at 10 µg

      Secondary
      Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

      Predicted band size: 64 kDa
      Observed band size: 74 kDa why is the actual band size different from the predicted?


      Exposure time: 3 minutes


      Blocking/Dilution buffer: 5% NFDM/TBST.

    • Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Western blot - Anti-Granulin antibody [EPR15864] (ab208777)
      Anti-Granulin antibody [EPR15864] (ab208777) at 1/1000 dilution + Human GRN full-length recombinant protein with GST-tag at N-terminal at 0.01 µg

      Secondary
      Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

      Predicted band size: 64 kDa
      Observed band size: 90 kDa why is the actual band size different from the predicted?


      Exposure time: 15 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)

      Immunohistochemical analysis of paraffin-embedded Human breast cancer tissue labeling Granulin with ab208777 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on Human breast cancer is observed. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)

      Immunohistochemical analysis of paraffin-embedded Human gastric cancer tissue labeling Granulin with ab208777 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on Human gastric cancer is observed. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Granulin antibody [EPR15864] (ab208777)

      Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling Granulin with ab208777 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on Kupffer cells of Human liver is observed. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    References

    Publishing research using ab208777? Please let us know so that we can cite the reference in this datasheet.

    ab208777 has not yet been referenced specifically in any publications.

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