Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-HADHA antibody [EPR17940] - BSA and Azide free (ab231169)

Overview

  • Product name

    Anti-HADHA antibody [EPR17940] - BSA and Azide free
    See all HADHA primary antibodies
  • Description

    Rabbit monoclonal [EPR17940] to HADHA - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human HADHA aa 600-700. The exact sequence is proprietary.
    Database link: P40939

  • Positive control

    • WB: HeLa, HEK-293, HepG2, Jurkat, C6 and RAW 264.7 cell lysates; Human fetal liver and fetal kidney lysates; Mouse heart and kidney lysates; Rat heart and kidney lysates. IHC-P: Human cervix carcinoma and tonsil tissues. ICC/IF: Jurkat and HeLa cells. Flow Cyt: HeLa cells. IP: HeLa whole cell lysate.
  • General notes

    Ab231169 is the carrier-free version of ab203114. 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.

    ab231169 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

Applications

Our Abpromise guarantee covers the use of ab231169 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 Use at an assay dependent concentration. Detects a band of approximately 74 kDa (predicted molecular weight: 83 kDa).
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG (Low endotoxin, Azide free), is suitable for use as an isotype control with this antibody.

 

IP Use at an assay dependent concentration.

Target

  • Function

    Bifunctional subunit.
  • Pathway

    Lipid metabolism; fatty acid beta-oxidation.
  • Involvement in disease

    Defects in HADHA are a cause of trifunctional protein deficiency (TFP deficiency) [MIM:609015]. The clinical manifestations are very variable and include hypoglycemia, cardiomyopathy and sudden death. Phenotypes with mainly hepatic and neuromyopathic involvement can also be distinguished. Biochemically, TFP deficiency is defined by the loss of all enzyme activities of the TFP complex.
    Defects in HADHA are the cause of long-chain 3-hydroxyl-CoA dehydrogenase deficiency (LCHAD deficiency) [MIM:609016]. The clinical features are very similar to TFP deficiency. Biochemically, LCHAD deficiency is characterized by reduced long-chain 3-hydroxyl-CoA dehydrogenase activity, while the other enzyme activities of the TFP complex are normal or only slightly reduced.
    Defects in HADHA are a cause of maternal acute fatty liver of pregnancy (AFLP) [MIM:609016]. AFLP is a severe maternal illness occurring during pregnancies with affected fetuses. This disease is associated with LCHAD deficiency and characterized by sudden unexplained infant death or hypoglycemia and abnormal liver enzymes (Reye-like syndrome).
  • Sequence similarities

    In the N-terminal section; belongs to the enoyl-CoA hydratase/isomerase family.
    In the central section; belongs to the 3-hydroxyacyl-CoA dehydrogenase family.
  • Cellular localization

    Mitochondrion.
  • Information by UniProt
  • Database links

  • Alternative names

    • 3 ketoacyl Coenzyme A (CoA) thiolase alpha subunit antibody
    • 3 oxoacyl CoA thiolase antibody
    • 78 kDa gastrin binding protein antibody
    • 78 kDa gastrin-binding protein antibody
    • ECHA antibody
    • ECHA_HUMAN antibody
    • GBP antibody
    • HADH antibody
    • HADHA antibody
    • Hydroxyacyl Coenzyme A dehydrogenase/3 ketoacyl Coenzyme A thiolase/enoyl Coenzyme A hydratase (trifunctional protein) alpha subunit antibody
    • LCEH antibody
    • LCHAD antibody
    • Long chain 3-hydroxyacyl-CoA dehydrogenase antibody
    • Mitochondrial long chain 2 enoyl Coenzyme A (CoA) hydratase alpha subunit antibody
    • Mitochondrial long chain L 3 hydroxyacyl Coenzyme A dehydrogenase alpha subunit antibody
    • Mitochondrial trifunctional enzyme alpha subunit antibody
    • Mitochondrial trifunctional protein alpha subunit antibody
    • MTPA antibody
    • Thiolase/enoyl Coenzyme A hydratase (trifunctional protein) alpha subunit antibody
    • TP ALPHA antibody
    • TP-alpha antibody
    • Trifunctional enzyme subunit alpha mitochondrial precursor antibody
    see all

Images

  • This WB data was generated using the same anti-HADHA antibody clone, EPR17940, in a different buffer formulation (cat# ab203114).

    Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: HADHA knockout HAP1 cell lysate (20 µg) 
    Lane 3: HEK293 cell lysate (20 µg)
    Lane 4: HepG2 cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab203114 observed at 82 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab203114 was shown to specifically react with HADHA when HADHA knockout samples were used. Wild-type and HADHA knockout samples were subjected to SDS-PAGE. ab203114 and ab8245 (loading control to GAPDH) were diluted at 1/1000 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 tonsil tissue labeling HADHA with ab203114 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasmic staining on Human tonsil tissue is observed. Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab203114).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Jurkat (Human T cell leukemia cells from peripheral blood) cells labeling HADHA with ab203114 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on Jurkat cell line.

    The nuclear counter stain is DAPI (blue). COX IV is detected with ab33985 (anti-COX IV Mitochondrial Marker mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab203114 at 1/250 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
    -ve control 2: ab33985 (anti-COX IV Mitochondrial Marker mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab203114).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling HADHA with ab203114 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HeLa cell line.

    The nuclear counter stain is DAPI (blue). COX IV is detected with ab33985 (anti-COX IV Mitochondrial Marker mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab203114 at 1/250 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
    -ve control 2: ab33985 (anti-COX IV Mitochondrial Marker mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab203114).

  • Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling HADHA with ab203114 at 1/100 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/500 dilution was used as the secondary antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab203114).

  • HADHA was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate with ab203114 at 1/400 dilution. Western blot was performed from the immunoprecipitate using ab203114 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.

    Lane 1: HeLa whole cell lysate 10ug (Input). Lane 2: ab203114 IP in HeLa whole cell lysate. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab203114 in HeLa whole cell lysate.

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

    Exposure time: 3 seconds.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab203114).

  • This IHC data was generated using the same anti-HADHA antibody clone, EPR17940, in a different buffer formulation (cat# ab203114).

    Immunohistochemical analysis of paraffin-embedded Human cervix carcinoma tissue labeling HADHA with ab203114 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasmic staining on Human cervix carcinoma tissue is observed. Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

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

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

References

ab231169 has not yet been referenced specifically in any publications.

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