Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Tissue Factor antibody [EPR22548-232] - BSA and Azide free (ab254010)

Overview

  • Product name

    Anti-Tissue Factor antibody [EPR22548-232] - BSA and Azide free
    See all Tissue Factor primary antibodies
  • Description

    Rabbit monoclonal [EPR22548-232] to Tissue Factor - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Human
  • Immunogen

    Recombinant fragment within Human Tissue Factor aa 1 to the C-terminus. The exact sequence is proprietary.
    Database link: P13726

  • Positive control

    • IHC-P: Human cervix carcinoma and kidney tissue. ICC/IF: A431 cells. Flow Cyt: A431 cells. IP: Tissue Factor IP in A431 whole cell lysate.
  • General notes

    Ab254010 is the carrier-free version of ab228968. 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.

    ab254010 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.

Properties

Applications

Our Abpromise guarantee covers the use of ab254010 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 1/1000. Detects a band of approximately 47 kDa (predicted molecular weight: 33 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.
IP Use at an assay dependent concentration.

Target

  • Function

    Initiates blood coagulation by forming a complex with circulating factor VII or VIIa. The [TF:VIIa] complex activates factors IX or X by specific limited protolysis. TF plays a role in normal hemostasis by initiating the cell-surface assembly and propagation of the coagulation protease cascade.
  • Sequence similarities

    Belongs to the tissue factor family.
  • Cellular localization

    Membrane.
  • Information by UniProt
  • Database links

  • Alternative names

    • CD142 antibody
    • CD142 antigen antibody
    • Coagulation factor III (thromboplastin tissue factor) antibody
    • Coagulation factor III antibody
    • F3 antibody
    • FLJ17960 antibody
    • TF antibody
    • TF_HUMAN antibody
    • TFA antibody
    • Thromboplastin antibody
    • Tissue factor antibody
    see all

Images

  • Immunohistochemical analysis of paraffin-embedded human kidney tissue labeling Tissue Factor with ab228968 at 1/500 dilution, followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining in human renal glomerulus (PMID: 7684196). The section was incubated with ab228968 for 30 mins at RT. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
    Perform heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab228968).

  • All lanes :

    Lane 1 : Wild-type HAP1 whole cell lysate at 60 µg
    Lane 2 : Tissue Factor knockout HAP1 whole cell lysate at 60 µg
    Lane 3 : A431 (human epidermoid carcinoma epithelial cell), whole cell lysate at 20 µg
    Lane 4 : MDA-MB-231 (human breast adenocarcinoma epithelial cell), whole cell lysate
    Lane 5 : MCF7 (human breast adenocarcinoma epithelial cell), whole cell lysate at 20 µg
    Lane 6 : BxPC-3 (human pancreas adenocarcinoma epithelial cell), whole cell lysate at 20 µg

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 33 kDa
    Observed band size: 47 kDa
    why is the actual band size different from the predicted?



    ab228968 was shown to specifically react with Tissue Factor in wild-type HAP1 cells as signal was lost in Tissue Factor knockout cells. Wild-type and Tissue Factor knockout samples were subjected to SDS-PAGE. ab228968 and ab181602 (Rabbit anti-GAPDH loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/200,000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument using the ECL technique.

    The molecular weight observed is consistent with what has been described in the literature (PMID: 28938620).

    Low expression: MCF-7 (PMID: 24137414, 28938620).

    Exposure times: Lanes 1-2: 70 secs; Lanes 3-6: 3.25 secs.

    Blocking/Dilution buffer: 5% NFDM/TBST.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab228968).

  • Immunohistochemical analysis of paraffin-embedded human cervix carcinoma tissue labeling Tissue Factor with ab228968 at 1/500 dilution, followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Membranous and weak cytoplasmic staining on tumor cells of human cervix carcinoma (PMID: 26383146). The section was incubated with ab228968 for 30 mins at RT. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Perform heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilizedA431 (human epidermoid carcinoma epithelial cell) and MCF7 () cells labeling Tissue Factor with ab228968 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous staining in A431 cell line. Low expression: MCF7 PMID: 24137414, 28938620. The nuclear stain is DAPI (blue).
    Counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at a 1/200 dilution (red).
    The negative control is the secondary antibody only.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab228968).

  • Flow cytometric analysis of MCF7 (human breast adenocarcinoma epithelial cell, Left) / A431 (human epidermoid carcinoma epithelial cell, Right) labeling Tissue Factor with ab228968 at 1/500 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

    Low expression: MCF7 (PMID: 24137414, PMID: 28938620).

    Gated on viable cells.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab228968).

  • Tissue Factor was immunoprecipitated from 0.35 mg A431 (human epidermoid carcinoma epithelial cell) whole cell lysate with ab228968 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab228968 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

    Lane 1: A431 whole cell lysate 10 µg (Input).
    Lane 2: ab228968 IP in A431 whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab228968 in A431 whole cell lysate.

    Blocking/Dilution buffer: 5% NFDM/TBST.
    Exposure time: 30 seconds.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab228968).

References

ab254010 has not yet been referenced specifically in any publications.

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