Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (ab224268)

Overview

  • Product name

    Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free
    See all Niemann Pick C1 primary antibodies
  • Description

    Rabbit monoclonal [EPR5209] to Niemann Pick C1 - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide aa 1250 to the C-terminus (C terminal). The exact sequence is proprietary.
    Database link: O15118

  • Positive control

    • 3T3L1 cell lysate, L6 cell lysate, HepG2 cell lysate, THP1 cell lysate, 293T cell lysate, PC3 cell lysate, Rat liver lysate, Rat brain lysate, Human kidney tissue.
  • General notes

    Ab224268 is the carrier-free version of ab134113. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab224268 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    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.

Applications

Our Abpromise guarantee covers the use of ab224268 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
ICC/IF Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 180 kDa (predicted molecular weight: 142 kDa).
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Target

  • Function

    Involved in the intracellular trafficking of cholesterol. May play a role in vesicular trafficking in glia, a process that may be crucial for maintaining the structural and functional integrity of nerve terminals.
  • Involvement in disease

    Defects in NPC1 are the cause of Niemann-Pick disease type C1 (NPDC1) [MIM:257220]. A lysosomal storage disorder that affects the viscera and the central nervous system. It is due to defective intracellular processing and transport of low-density lipoprotein derived cholesterol. It causes accumulation of cholesterol in lysosomes, with delayed induction of cholesterol homeostatic reactions. Niemann-Pick disease type C1 has a highly variable clinical phenotype. Clinical features include variable hepatosplenomegaly and severe progressive neurological dysfunction such as ataxia, dystonia and dementia. The age of onset can vary from infancy to late adulthood. An allelic variant of Niemann-Pick disease type C1 is found in people with Nova Scotia ancestry. Patients with the Nova Scotian clinical variant are less severely affected.
  • Sequence similarities

    Belongs to the patched family.
    Contains 1 SSD (sterol-sensing) domain.
  • Domain

    A cysteine-rich N-terminal domain and a C-terminal domain containing a di-leucine motif necessary for lysosomal targeting are critical for mobilization of cholesterol from lysosomes.
  • Post-translational
    modifications

    Glycosylated.
  • Cellular localization

    Late endosome membrane. Lysosome membrane.
  • Information by UniProt
  • Database links

  • Alternative names

    • Niemann Pick C1 protein precursor antibody
    • Niemann Pick disease, type C1 antibody
    • Niemann-Pick C1 protein antibody
    • NPC antibody
    • NPC1 antibody
    • NPC1_HUMAN antibody
    see all

Images

  • All lanes : Anti-Niemann Pick C1 antibody [EPR5209] (ab134113) at 1/1000 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : NPC1 (Niemann Pick C1) knockout HAP1 whole cell lysate
    Lane 3 : HEK293 whole cell lysate
    Lane 4 : HepG2 whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 142 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab134113 observed at 180 kDa. Red - loading control, ab9484, observed at 37 kDa.

    ab134113 was shown to specifically react with Niemann Pick C1 in wild-type HAP1 cells as signal was lost in NPC1 (Niemann Pick C1) knockout cells. Wild-type and NPC1 (Niemann Pick C1) knockout samples were subjected to SDS-PAGE. Ab134113 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113). 

  • Cathepsin B/L inhibition causes NPC disease-like cholesterol accumulation in SH-SY5Y cells.

    Confocal microscopy of SH-SY5Y control and PADK treated cells. Cholesterol (filipin staining, white) and NPC1 (ab134113; green).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • ab134113 staining Niemann Pick C1 in Neuro-2a (mouse neuroblastoma cell line) by flow cytometry. Cells were fixed with 4% paraformaldehyde, permabilised with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/200. A goat anti rabbit IgG (Alexa Fluor® 488) at a dilution of 1/2000 was used as the secondary antibody.

    Isoytype control: Rabbit monoclonal IgG (Black)

    Unlabeled control: Cell without incubation with primary antibody and secondary antibody (Blue)

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • Immunofluorescence staining of Neuro-2a (mouse neuroblastoma cell line) cells with purified ab134113 at a working dilution of 1 in 70, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti rabbit (ab150077), used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, purified ab134113 was used at a dilution of 1/200 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • Immunohistochemical staining of paraffin embedded rat cerebellum with purified ab134113 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. 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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • Immunohistochemical staining of paraffin embedded mouse liver with purified ab134113 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. 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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • Immunohistochemical staining of paraffin embedded human liver with purified ab134113 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. 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.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • Immunohistochemical analysis of paraffin embedded human kidney tissue labelling Niemann Pick C1 with unpurified ab134113 at 1/50.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

    Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

  • Equilibrium disassociation constant (KD)
    Learn more about KD

    Click here to learn more about KD

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

  • This ICC data was generated using the same anti-Niemann Pick C1 antibody clone [EPR5209] in a different buffer formulation (cat# ab134133).

    Immunofluorescence staining of neuro-2a cells with purified ab134113 at a working dilution of 1 in 70, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti rabbit (ab150077), used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, purified ab134113 was used at a dilution of 1/200 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500.

References

This product has been referenced in:

  • Cockburn CL  et al. Functional inhibition of acid sphingomyelinase disrupts infection by intracellular bacterial pathogens. Life Sci Alliance 2:N/A (2019). Read more (PubMed: 30902833) »
See 1 Publication for this product

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