Validated using a knockout cell line
Recombinant
RabMAb

Anti-PFKFB3 antibody [EPR12594] (ab181861)

Knockout Tested Rabbit recombinant monoclonal PFKFB3 antibody [EPR12594]. Validated in WB, IP, IHC, Flow Cyt, ICC/IF and tested in Mouse, Rat, Human. Cited in 4 publication(s).

Overview

  • Product name
    Anti-PFKFB3 antibody [EPR12594]
    See all PFKFB3 primary antibodies
  • Description
    Rabbit monoclonal [EPR12594] to PFKFB3
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, IP, ICC/IF, IHC-P, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Recombinant fragment within Human PFKFB3 aa 300 to the C-terminus. The exact sequence is proprietary.
    Database link: Q16875

  • Positive control
    • Jurkat and HeLa whole cell lysate (ab150035); Human melanoma tissue; HeLa and A431 cells.
  • 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.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab181861 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 - 1/10000. Detects a band of approximately 58 kDa (predicted molecular weight: 60 kDa).
IP 1/50.
ICC/IF 1/100.
IHC-P 1/50.
Flow Cyt Use at an assay dependent concentration.

Target

  • Function
    Synthesis and degradation of fructose 2,6-bisphosphate.
  • Tissue specificity
    Ubiquitous.
  • Sequence similarities
    In the C-terminal section; belongs to the phosphoglycerate mutase family.
  • Post-translational
    modifications
    Phosphorylation by AMPK stimulates activity.
  • Information by UniProt
  • Database links
  • Alternative names
    • 6 phosphofructo 2 kinase/ fructose 2,6 bisphosphatase antibody
    • 6 phosphofructo 2 kinase/fructose 2,6 biphosphatase 3 antibody
    • 6-bisphosphatase antibody
    • 6-P2ase 3 antibody
    • 6-P2ASE brain/placenta-type isozyme antibody
    • 6PF 2 K/Fru 2,6 P2ASE brain/placenta type isozyme antibody
    • 6PF 2-K/Fru 2,6 P2ase 3 antibody
    • 6PF-2-K/Fru-2 antibody
    • F263_HUMAN antibody
    • fructose 6 phosphate,2 kinase/fructose 2, 6 bisphosphatase antibody
    • Fructose-2 antibody
    • Inducible 6 phosphofructo 2 kinase/fructose 2,6 bisphosphatase antibody
    • iPFK 2 antibody
    • iPFK-2 antibody
    • IPFK2 antibody
    • PFK/FBPase 3 antibody
    • PFK2 antibody
    • PFKFB3 antibody
    • Renal carcinoma antigen NY REN 56 antibody
    • Renal carcinoma antigen NY-REN-56 antibody
    • uPFK antibody
    see all

Images

  • Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: PFKFB3 knockout HAP1 whole cell lysate (20 µg)
    Lane 3: HeLa whole cell lysate (20 µg)
    Lane 4: Jurkat whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab181861 observed at 60 kDa. Red - loading control, ab9484, observed at 37 kDa.

    ab181861 was shown to specifically react with PFKFB3 in wild-type HAP1 cells as signal was lost in PFKFB3 knockout cells. Wild-type and PFKFB3 knockout samples were subjected to SDS-PAGE. Ab181861 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.

  • Immunofluorescent analysis of acetone-fixed HeLa cells labeling PFKFB3 with ab181861 at 1/100 dilution, followed by Goat anti rabbit IgG (Alexa Fluor®555) at 1/200 dilution. Counter stained with Dapi (blue).

  • All lanes : Anti-PFKFB3 antibody [EPR12594] (ab181861) at 1/20000 dilution

    Lane 1 : Jurkat cell lysate
    Lane 2 : HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugate at 1/1000 dilution

    Predicted band size: 60 kDa
    Observed band size: 58 kDa
    why is the actual band size different from the predicted?



    Blocking buffer:  5% NFDM/TBST

  • Immunohistochemical analysis of paraffin-embedded human melanoma tissue labeling PFKFB3 with ab181861 at 1/50 dilution followed by prediluted HRP Polymer for Rabbit IgG. Counter stained with Hematoxylin.

  • Flow Cytometry analysis of A431(human epidermoid carcinoma) cells labeling PFKFB3 with purified ab181861 at 1/210 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488)(1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) was used as the unlabeled control.

  • Western blot analysis of PFKFB3 in HeLa cell lysate immunoprecipitated using ab181861 at 1/50 dilution (Lane 1). Lane 2: Negative control.

    Secondary antibody: Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1500 dilution.

     

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed A431 cells labeling PFKFB3 with ab181861 at 1/100 dilution, followed by Goat anti rabbit IgG (Alexa Fluor®555) at 1/200 dilution. Counter stained with Dapi (blue).

References

This product has been referenced in:
  • Xu C  et al. Canagliflozin exerts anti-inflammatory effects by inhibiting intracellular glucose metabolism and promoting autophagy in immune cells. Biochem Pharmacol 152:45-59 (2018). WB ; Mouse . Read more (PubMed: 29551587) »
  • Yan S  et al. 6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoform 3 spatially mediates autophagy through the AMPK signaling pathway. Oncotarget 8:80909-80922 (2017). Read more (PubMed: 29113354) »
See all 4 Publications for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Application
Western blot
Sample
Cow Cell lysate - whole cell (VVEC)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
20 µg
Treatment
100 uM ATP 0, 5 min, 30 min
Specification
VVEC
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Jun 03 2016

Application
Western blot
Sample
Mouse Cell lysate - whole cell (primary alveolar type II cells)
Gel Running Conditions
Non-reduced Denaturing (10%)
Loading amount
20 µg
Specification
primary alveolar type II cells
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Nov 02 2015

Application
Western blot
Sample
Human Cell lysate - whole cell (A549 cells)
Gel Running Conditions
Non-reduced Denaturing (10)
Loading amount
20 µg
Specification
A549 cells
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Dr. Christine Vohwinkel

Verified customer

Submitted Oct 29 2015

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

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