Overview

  • Product name
    Anti-Ndufs4 antibody [EP7832]
    See all Ndufs4 primary antibodies
  • Description
    Rabbit monoclonal [EP7832] to Ndufs4
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, IHC-P, ICC/IF, IP, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human Ndufs4 aa 150-250 (C terminal). The exact sequence is proprietary.

  • Positive control
    • 293T cell lysates, fetal brain and fetal stomach tissue lysates; Human brain and Human stomach tissues Mouse heat lysate, rat heart lysate, mouse kidney, HeLa.
  • 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.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab137064 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 1/500 - 1/10000. Detects a band of approximately 18 kDa (predicted molecular weight: 20 kDa).
IHC-P 1/50 - 1/100. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ICC/IF 1/500.

For unpurified use at 1/50 - 1/100. 

IP 1/10 - 1/100.
Flow Cyt 1/60.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

For unpurifed use at 1/100 - 1/1000 dilution. 

 

Target

  • Function
    Accessory subunit of the mitochondrial membrane respiratory chain NADH dehydrogenase (Complex I), that is believed not to be involved in catalysis. Complex I functions in the transfer of electrons from NADH to the respiratory chain. The immediate electron acceptor for the enzyme is believed to be ubiquinone.
  • Involvement in disease
    Defects in NDUFS4 are a cause of mitochondrial complex I deficiency (MT-C1D) [MIM:252010]. A disorder of the mitochondrial respiratory chain that causes a wide range of clinical disorders, from lethal neonatal disease to adult-onset neurodegenerative disorders. Phenotypes include macrocephaly with progressive leukodystrophy, non-specific encephalopathy, cardiomyopathy, myopathy, liver disease, Leigh syndrome, Leber hereditary optic neuropathy, and some forms of Parkinson disease.
  • Sequence similarities
    Belongs to the complex I NDUFS4 subunit family.
  • Cellular localization
    Mitochondrion inner membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • AQDQ antibody
    • CI 18 antibody
    • CI 18 kDa antibody
    • CI AQDQ antibody
    • CI-18 kDa antibody
    • CI-AQDQ antibody
    • Complex I 18 kDa antibody
    • Complex I AQDQ antibody
    • Complex I-18 kDa antibody
    • Complex I-AQDQ antibody
    • mitochondrial antibody
    • mitochondrial respiratory chain complex I (18 KD subunit) antibody
    • NADH coenzyme Q reductase antibody
    • NADH dehydrogenase (ubiquinone) Fe S protein 4 18kDa antibody
    • NADH dehydrogenase [ubiquinone] iron-sulfur protein 4 antibody
    • NADH dehydrogenase antibody
    • NADH ubiquinone oxidoreductase 18 kDa subunit antibody
    • NADH-ubiquinone oxidoreductase 18 kDa subunit antibody
    • NDUFS4 antibody
    • NDUS4_HUMAN antibody
    see all

Images

  • Anti-Ndufs4 antibody [EP7832] (ab137064) at 1/50000 dilution (purified) + Rat heart lysates at 15 µg

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

    Predicted band size: 20 kDa



    Blocking and diluting buffer: 5% NFDM/TBST

  • ab137064 (purified) at 1:30 dilution (2μg) immunoprecipitating Ndufs4 in Rat heart lysate.

    Lane 1 (input): Rat heart lysate, 10μg
    Lane 2 (+): ab137064 & Rat heart lysate
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab137064 in Rat heart lysate

    For western blotting, VeriBlot for IP secondary antibody (HRP) (ab131366) was used as the secondary antibody at 1:1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

  • Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Ndufs4 with purified ab137064 at 1:60 dilution (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor ® 488) secondary antibody was used at 1:2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).
  • Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Ndusf5 with Purified ab137064 at 1:500 dilution. Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab7291 anti-Tubulin (mouse mAb) ab150120 AlexaFluor ® 594 Goat anti-Mouse secondary (1:1000,2 μg/ml). ab150077 Goat anti rabbit IgG(Alexa Fluor ® 488) was used as the secondary antibody at 1:1000 dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat cerebrum tissue sections labeling Ndufs4 with Purified ab137064 at 1:50 dilution. Heat mediated antigen retrieval was performed using Tris/EDTA buffer, pH 9.0. Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse kidney tissue sections labeling Ndufs4 with Purified ab137064 at 1:50 dilution. Heat mediated antigen retrieval was performed using Tris/EDTA buffer, pH 9.0. Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human stomach tissue sections labeling Ndufs4 with Purified ab137064 at 1:50 dilution. Heat mediated antigen retrieval was performed using Tris/EDTA buffer, pH 9.0. Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
  • All lanes : Anti-Ndufs4 antibody [EP7832] (ab137064) at 1/2000 dilution (purified)

    Lane 1 : Human fetal brain lysates
    Lane 2 : Mouse heart lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

    Predicted band size: 20 kDa
    Observed band size: 18 kDa
    why is the actual band size different from the predicted?



    Blocking and diluting buffer: 5% NFDM/TBST.

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: Ndufs4 knockout HAP1 cell lysate (20 µg)
    Lane 3: HEK293 cell lysate (20 µg)
    Lane 4: Human fetal heart tissue lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab137064 observed at 23 kDa. Red - loading control, ab8245, observed at 37 kDa.

    Unpurified ab137064 was shown to recognize Ndufs4 when Ndufs4 knockout samples were used, along with additional cross-reactive bands. Wild-type and Ndufs4 knockout samples were subjected to SDS-PAGE. ab137064 and ab8245 (loading control to GAPDH) were diluted at 1/500 and 1/10 000 respectively 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 1/10000 dilution for 1 h at room temperature before imaging.

  • Immunohistochemical analysis of paraffin-embedded Human brain tissue labelling Ndufs4 with unpurified ab137064 at 1/50 dilution.

  • Overlay histogram showing HepG2 cells stained with unpurified ab137064 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab137064, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in HepG2 cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

  • All lanes : Anti-Ndufs4 antibody [EP7832] (ab137064) at 1/1000 dilution (unpurified)

    Lane 1 : 293T cell lysate
    Lane 2 : Fetal brain tissue lysate
    Lane 3 : Fetal kidney tissue lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 20 kDa
    Observed band size: 18 kDa why is the actual band size different from the predicted?

  • Immunohistochemical analysis of paraffin-embedded Human stomach tissue labelling Ndufs4 with unpurified ab137064 at 1/50 dilution.

References

This product has been referenced in:
  • Piekutowska-Abramczuk D  et al. NDUFB8 Mutations Cause Mitochondrial Complex I Deficiency in Individuals with Leigh-like Encephalomyopathy. Am J Hum Genet 102:460-467 (2018). Read more (PubMed: 29429571) »
See 1 Publication for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Kidney)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization
No
Specification
Kidney
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 21°C
Fixative
Formaldehyde

Mr. Carl Hobbs

Verified customer

Submitted Feb 11 2019

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Kidney)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization
No
Specification
Kidney
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 21°C
Fixative
Formaldehyde

Mr. Carl Hobbs

Verified customer

Submitted Feb 11 2019

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Rat Tissue sections (Kidney)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization
No
Specification
Kidney
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 21°C
Fixative
Formaldehyde

Mr. Carl Hobbs

Verified customer

Submitted Feb 11 2019

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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