Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)

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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 [EPR15010] to Sortilin/NT3
  • Suitable for: IHC-P, WB
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-Sortilin/NT3 antibody [EPR15010]
    See all Sortilin/NT3 primary antibodies

  • Description

    Rabbit monoclonal [EPR15010] to Sortilin/NT3

  • Host species

    Rabbit

  • Tested applications

    Suitable for: IHC-P, WBmore details

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant fragment within Human Sortilin/NT3 aa 650-800. The exact sequence is proprietary.
    Database link: Q99523

  • Positive control

    • WB: SW480 whole cell lysate (ab3957), SW40, HeLa and SH-SY5Y cell lysates. IHC-P: Human brain tissues.

  • 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 ab188586 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
IHC-P 1/250. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB 1/1000 - 1/2000. Detects a band of approximately 92-114 kDa (predicted molecular weight: 92 kDa).

Target

  • Function

    Functions as a sorting receptor in the Golgi compartment and as a clearance receptor on the cell surface. Required for protein transport from the Golgi apparatus to the lysosomes by a pathway that is independent of the mannose-6-phosphate receptor (M6PR). Also required for protein transport from the Golgi apparatus to the endosomes. Promotes neuronal apoptosis by mediating endocytosis of the proapoptotic precursor forms of BDNF (proBDNF) and NGFB (proNGFB). Also acts as a receptor for neurotensin. May promote mineralization of the extracellular matrix during osteogenic differentiation by scavenging extracellular LPL. Probably required in adipocytes for the formation of specialized storage vesicles containing the glucose transporter SLC2A4/GLUT4 (GLUT4 storage vesicles, or GSVs). These vesicles provide a stable pool of SLC2A4 and confer increased responsiveness to insulin. May also mediate transport from the endoplasmic reticulum to the Golgi.

  • Tissue specificity

    Expressed at high levels in brain, spinal cord, heart, skeletal muscle, thyroid, placenta and testis. Expressed at lower levels in lymphoid organs, kidney, colon and liver.

  • Involvement in disease

    Note=A common polymorphism located in a non-coding region between CELSR2 and PSRC1 alters a CEBP transcription factor binding site and is responsible for changes in hepatic expression of SORT1. Altered SORT1 expression in liver affects low density lipoprotein cholesterol levels in plasma and is associated with susceptibility to myocardial infarction.

  • Sequence similarities

    Belongs to the VPS10-related sortilin family. SORT1 subfamily.
    Contains 9 BNR repeats.

  • Domain

    The N-terminal propeptide may facilitate precursor transport within the Golgi stack. Intrachain binding of the N-terminal propeptide and the extracellular domain may also inhibit premature ligand binding.
    The extracellular domain may be shed following protease cleavage in some cell types.

  • Post-translational
    modifications

    The N-terminal propeptide is cleaved by furin and possibly other homologous proteases.

  • Cellular localization

    Membrane. Endoplasmic reticulum membrane. Endosome membrane. Golgi apparatus > Golgi stack membrane. Lysosome membrane. Nucleus membrane. Cell membrane. Lysosome membrane. Localized to membranes of the endoplasmic reticulum, endosomes, Golgi stack, lysosomes and nucleus. A small fraction of the protein is also localized to the plasma membrane. May also be found in SLC2A4/GLUT4 storage vesicles (GSVs) in adipocytes. Localization to the plasma membrane in adipocytes may be enhanced by insulin.
  • Information by UniProt

  • Database links

    • Alternative names

      • 100 kDa NT receptor antibody
      • Glycoprotein 95 antibody
      • Gp 95 antibody
      • Gp95 antibody
      • LDLCQ6 antibody
      • Neurotensin receptor 3 antibody
      • NT 3 antibody
      • NT3 antibody
      • NTR 3 antibody
      • NTR3 antibody
      • OTTHUMP00000013784 antibody
      • SORT 1 antibody
      • SORT_HUMAN antibody
      • SORT1 (gene name) antibody
      • Sort1 antibody
      • Sortilin 1 antibody
      • Sortilin antibody
      see all

    Images

    • Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      All lanes : Anti-Sortilin/NT3 antibody [EPR15010] (ab188586) at 1/1000 dilution

      Lane 1 : Wild-type HeLa cell lysate
      Lane 2 : SORT1 knockout HeLa cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 92 kDa
      Observed band size: 100 kDa
      why is the actual band size different from the predicted?



      Lanes 1- 2: Merged signal (red and green). Green - ab188586 observed at 100 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

       ab188586 was shown to react with Sortilin/NT3 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264772 (knockout cell lysate ab257696) was used. Wild-type HeLa and SORT1 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab188586 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at a 1 in 1000 dilution and a 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-Sortilin/NT3 antibody [EPR15010] (ab188586)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)

      Immunohistochemical analysis of paraffin-embedded Human brain tissue labeling Sortilin/NT3 with ab188586 at 1/250 dilution followed by prediluted HRP Polymer for Rabbit IgG. Counter stained with Hematoxylin.

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

    • Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)

      Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: Sortilin/NT3 knockout HAP1 cell lysate (20 µg)
      Lane 3: SW480 cell lysate (20 µg)
      Lane 4: HeLa cell lysate (20 µg)

      Lanes 1 - 4: Merged signal (red and green). Green - ab188586 observed at 92 kDa. Red - loading control, ab8245, observed at 37 kDa.


      ab188586 was shown to  react with Sortilin/NT3 in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when Sortilin/NT3 knockout samples were examined. Wild-type and Sortilin/NT3 knockout samples were subjected to SDS-PAGE. ab188586 and ab8245 (loading control to GAPDH) were both diluted 1/2000 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.

    • Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      All lanes : Anti-Sortilin/NT3 antibody [EPR15010] (ab188586) at 1/1000 dilution

      Lane 1 : SH-SY5Y whole cell lysate
      Lane 2 : Human hippocampus tissue lysate
      Lane 3 : Human fetal brain tissue lysate
      Lane 4 : Mouse brain tissue lysate
      Lane 5 : 3T3-L1 whole cell lysate
      Lane 6 : Rat brain tissue lysate

      Lysates/proteins at 15 µg per lane.

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

      Predicted band size: 92 kDa
      Observed band size: 92 kDa


      Exposure time: 30 seconds


      Blocking and dilution buffer: 5% NFDM/TBST.

    • Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      Western blot - Anti-Sortilin/NT3 antibody [EPR15010] (ab188586)
      All lanes : Anti-Sortilin/NT3 antibody [EPR15010] (ab188586) at 1/2000 dilution

      Lane 1 : SW480 cell lysate
      Lane 2 : HeLa cell lysate
      Lane 3 : SH-SY5Y cell lysate

      Lysates/proteins at 10 µg per lane.

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

      Predicted band size: 92 kDa

    References

    Publishing research using ab188586? Please let us know so that we can cite the reference in this datasheet.

    ab188586 has been referenced in 1 publication.

    • Hirst J  et al. Role of the AP-5 adaptor protein complex in late endosome-to-Golgi retrieval. PLoS Biol 16:e2004411 (2018). PubMed: 29381698

    Customer reviews and Q&As

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    IHC - Wholemount abreview for Anti-Sortilin/NT3 antibody [EPR15010]

     Poor
    Abreviews
    Application
    IHC - Wholemount
    Sample
    Mouse Tissue (Coronal mouse brain sections)
    Specification
    Coronal mouse brain sections
    Read More

    Swetha Gowrishankar

    Verified customer

    Submitted May 16 2019

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