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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-DIAPH1 antibody [EPR7948] (ab129167)

Reviews (1) Submit a question References (7)
Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Immunocytochemistry/ Immunofluorescence - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Flow Cytometry (Intracellular) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
  • Anti-DIAPH1 antibody [EPR7948] (ab129167)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR7948] to DIAPH1
  • Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IF
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Alexa Fluor® 647 APC Carrier Free HRP PE

Overview

  • Product name

    Anti-DIAPH1 antibody [EPR7948]
    See all DIAPH1 primary antibodies

  • Description

    Rabbit monoclonal [EPR7948] to DIAPH1

  • Host species

    Rabbit

  • Tested applications

    Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IFmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide within Human DIAPH1 aa 1-100. The exact sequence is proprietary.
    Database link: O60610

  • Positive control

    • WB: HCT116 and 293T cell lysates' HEK-293, RAW 264.7 and Human brain lysates, PC-12 and MCF7 lysates; Flow Cyt (intra): HeLa cells; ICC/IF: HeLa cells; IHC-P: Human, rat, and mouse kidney tissue sections.

  • 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.

    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.

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.

  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
  • Concentration information loading...

  • Purity

    Protein A purified

  • Clonality

    Monoclonal

  • Clone number

    EPR7948

  • Isotype

    IgG

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab129167 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
Flow Cyt (Intra)
1/50.
WB (1)
1/1000 - 1/10000. Detects a band of approximately 150 kDa (predicted molecular weight: 141 kDa).
IHC-P
1/50 - 1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.
ICC/IF
1/50.

For unpurified use at 1/100 - 1/250.
Notes
Flow Cyt (Intra)
1/50.
WB
1/1000 - 1/10000. Detects a band of approximately 150 kDa (predicted molecular weight: 141 kDa).
IHC-P
1/50 - 1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.
ICC/IF
1/50.

For unpurified use at 1/100 - 1/250.

Target

  • Function

    Acts in a Rho-dependent manner to recruit PFY1 to the membrane. Required for the assembly of F-actin structures, such as actin cables and stress fibers. Nucleates actin filaments. Binds to the barbed end of the actin filament and slows down actin polymerization and depolymerization. Required for cytokinesis, and transcriptional activation of the serum response factor. DFR proteins couple Rho and Src tyrosine kinase during signaling and the regulation of actin dynamics. Functions as a scaffold protein for MAPRE1 and APC to stabilize microtubules and promote cell migration (By similarity). Has neurite outgrowth promoting activity (By similarity). In hear cells, it may play a role in the regulation of actin polymerization in hair cells. The MEMO1-RHOA-DIAPH1 signaling pathway plays an important role in ERBB2-dependent stabilization of microtubules at the cell cortex. It controls the localization of APC and CLASP2 to the cell membrane, via the regulation of GSK3B activity. In turn, membrane-bound APC allows the localization of the MACF1 to the cell membrane, which is required for microtubule capture and stabilization.

  • Tissue specificity

    Expressed in brain, heart, placenta, lung, kidney, pancreas, liver, skeletal muscle and cochlea.

  • Involvement in disease

    Defects in DIAPH1 are the cause of deafness autosomal dominant type 1 (DFNA1) [MIM:124900]. DFNA1 is a form of sensorineural hearing loss. Sensorineural deafness results from damage to the neural receptors of the inner ear, the nerve pathways to the brain, or the area of the brain that receives sound information.

  • Sequence similarities

    Belongs to the formin homology family. Diaphanous subfamily.
    Contains 1 DAD (diaphanous autoregulatory) domain.
    Contains 1 FH1 (formin homology 1) domain.
    Contains 1 FH2 (formin homology 2) domain.
    Contains 1 GBD/FH3 (Rho GTPase-binding/formin homology 3) domain.

  • Domain

    DRFs are regulated by intramolecular GBD-DAD binding where Rho-GTP activates the DRFs by disrupting the GBD-DAD interaction (By similarity). DCAF7 binds to the FH2 (formin homology 2) domain.

  • Cellular localization

    Cell membrane. Cell projection > ruffle membrane. Cytoplasm > cytoskeleton. Membrane ruffles, especially at the tip of ruffles, of motile cells.
  • Information by UniProt

  • Database links

    • Alternative names

      • DIAPH1 antibody
      • deafness, autosomal dominant 1 antibody
      • DFNA1 antibody
      • DIA1 antibody
      • DIAP1 antibody
      • DIAP1_HUMAN antibody
      • DIAPH1 antibody
      • Diaphanous homolog 1 (Drosophila) antibody
      • diaphanous homolog 1 antibody
      • Diaphanous related formin 1 antibody
      • Diaphanous-related formin-1 antibody
      • DRF1 antibody
      • FLJ25265 antibody
      • hDIA1 antibody
      • LFHL1 antibody
      • low frequency hearing loss 1 antibody
      • p140DIA antibody
      • Protein diaphanous homolog 1 antibody
      see all

    Images

    • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      All lanes : Anti-DIAPH1 antibody [EPR7948] (ab129167) at 1/10000 dilution (Purified)

      Lane 1 : PC-12 (Rat adrenal gland pheochromocytoma ) whole cell lysate
      Lane 2 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate

      Lysates/proteins at 20 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: 141 kDa

    • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      All lanes : Anti-DIAPH1 antibody [EPR7948] (ab129167) at 1/1000 dilution (Purified)

      Lane 1 : HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate
      Lane 2 : Human brain lysate
      Lane 3 : RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate

      Lysates/proteins at 20 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: 141 kDa

    • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      All lanes : Anti-DIAPH1 antibody [EPR7948] (ab129167) at 1/1000 dilution

      Lane 1 : Wild-type HEK-293T cell lysate
      Lane 2 : DIAPH1 knockout HEK-293T cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 141 kDa
      Observed band size: 150 kDa why is the actual band size different from the predicted?



      Lanes 1- 2: Merged signal (red and green). Green - ab129167 observed at 150 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.

       ab129167 was shown to react with DIAPH1 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266120 (knockout cell lysate ab257411) was used. Wild-type HEK-293T and DIAPH1 knockout HEK-293T 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. ab129167 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) 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.

    • Immunocytochemistry/ Immunofluorescence - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunocytochemistry/ Immunofluorescence - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunocytochemistry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling DIAPH1 with Purified ab129167 at 1/50 dilution (10 µg/mL). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 dilution (2.5 µg/mL). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 dilution (2 µg/mL). DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney tissue sections labeling DIAPH1 with Purified ab129167 at 1/100 dilution (5.96 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse kidney tissue sections labeling DIAPH1 with Purified ab129167 at 1/100 dilution (5.96 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue sections labeling DIAPH1 with Purified ab129167 at 1/100 dilution (5.96 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
    • Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Western blot - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      All lanes : Anti-DIAPH1 antibody [EPR7948] (ab129167) at 1/1000 dilution

      Lane 1 : Wild-type HCT116 cell lysate
      Lane 2 : DIAPH1 knockout HCT116 cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 141 kDa
      Observed band size: 155 kDa why is the actual band size different from the predicted?



      Lanes 1 - 2: Merged signal (red and green). Green - ab129167 observed at 155 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

      ab129167 was shown to react with DIAPH1 in HCT 116 wild-type cells in western blot with loss of signal observed in DIAPH1 knockout cell line ab273727 (DIAPH1 knockout cell lysate ab275252). HCT 116 wild-type and DIAPH1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab129167 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 hour at room temperature before imaging.

    • Flow Cytometry (Intracellular) - Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Flow Cytometry (Intracellular) - Anti-DIAPH1 antibody [EPR7948] (ab129167)

      Intracellular Flow Cytometry analysis of HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling DIAPH1 with purified ab129167 at 1/50 dilution. Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 was used as the secondary antibody (red). Rabbit monoclonal IgG (ab172730) was used as the isotype control (black). Cells without incubation with primary and secondary antibodies were used as the unlabeled control (blue). 

       

       

    • Anti-DIAPH1 antibody [EPR7948] (ab129167)
      Anti-DIAPH1 antibody [EPR7948] (ab129167)

    References (7)

    Publishing research using ab129167? Please let us know so that we can cite the reference in this datasheet.

    ab129167 has been referenced in 7 publications.

    • Lehtimäki JI  et al. Generation of stress fibers through myosin-driven reorganization of the actin cortex. Elife 10:N/A (2021). PubMed: 33506761
    • Zhang W  et al. Formin Activity and mDia1 Contribute to Maintain Axon Initial Segment Composition and Structure. Mol Neurobiol 58:6153-6169 (2021). PubMed: 34458961
    • Wu JH  et al. Inhibition of Sema4D/PlexinB1 signaling alleviates vascular dysfunction in diabetic retinopathy. EMBO Mol Med 12:e10154 (2020). PubMed: 31943789
    • Rebowski G  et al. Mechanism of actin N-terminal acetylation. Sci Adv 6:eaay8793 (2020). PubMed: 32284999
    • Shinozuka K  et al. Empathy in stroke rats is modulated by social settings. J Cereb Blood Flow Metab 40:1182-1192 (2020). PubMed: 31366299
    • Yang J  et al. DIAPH1 Is Upregulated and Inhibits Cell Apoptosis through ATR/p53/Caspase-3 Signaling Pathway in Laryngeal Squamous Cell Carcinoma. Dis Markers 2019:6716472 (2019). PubMed: 30733838
    • Jeong J  et al. Soluble RAGE attenuates AngII-induced endothelial hyperpermeability by disrupting HMGB1-mediated crosstalk between AT1R and RAGE. Exp Mol Med 51:1-15 (2019). PubMed: 31562296

    Customer reviews and Q&As

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    Western blot abreview for Anti-DIAPH1 antibody [EPR7948]

     Excellent
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (HeLa)
    Gel Running Conditions
    Reduced Denaturing (4-12% gradient Bis Tris gel ran 1X MOPS)
    Loading amount
    10 µg
    Specification
    HeLa
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

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    Verified customer

    Submitted Nov 05 2018

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