Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-FHL2 antibody [EPR17860-23] (ab202586)

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Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR17860-23] to FHL2
  • Suitable for: WB, ICC/IF, IP
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-FHL2 antibody [EPR17860-23]
    See all FHL2 primary antibodies

  • Description

    Rabbit monoclonal [EPR17860-23] to FHL2

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, ICC/IF, IPmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Recombinant fragment within Human FHL2 aa 1 to the C-terminus. The exact sequence is proprietary.
    Database link: Q14192

  • Positive control

    • WB: K562, SW480, PC-12, HT1080 and HeLa whole cell lysates; Human fetal heart, Mouse heart, mouse kidney, rat heart and rat kidney lysates. ICC/IF: A-673 and NIH/3T3 cells. IP: SW480 whole cell lysate. ICC/IF: U2OS cells (U2OS-FHL2 cell line used as a negative control).

  • 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.

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

    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...

  • Purity

    Protein A purified

  • Clonality

    Monoclonal

  • Clone number

    EPR17860-23

  • Isotype

    IgG

  • Research areas

Applications

Our Abpromise guarantee covers the use of ab202586 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 32 kDa (predicted molecular weight: 32 kDa).
ICC/IF 1/100.

ab202586 works both with PFA and methanol fixation.
IP 1/20.

Target

  • Function

    May function as a molecular transmitter linking various signaling pathways to transcriptional regulation. Negatively regulates the transcriptional repressor E4F1 and may function in cell growth.

  • Tissue specificity

    Expressed in skeletal muscle and heart.

  • Sequence similarities

    Contains 4 LIM zinc-binding domains.

  • Domain

    The third LIM zinc-binding mediates interaction with E4F1.

  • Cellular localization

    Cytoplasm. Nucleus.
  • Information by UniProt

  • Database links

    see all

  • Alternative names

    • AAG 11 antibody
    • AAG11 antibody
    • Aging associated gene 11 antibody
    • Down regulated in rhabdomyosarcoma LIM protein antibody
    • Downregulated in rhabdomyosarcoma LIM protein antibody
    • DRAL antibody
    • FHL 2 antibody
    • FHL-2 antibody
    • Fhl2 antibody
    • FHL2 protein antibody
    • FHL2_HUMAN antibody
    • Four and a half LIM domain protein 2 antibody
    • Four and a half LIM domains 2 antibody
    • Four and a half LIM domains protein 2 antibody
    • KIAA0990 antibody
    • LIM domain protein DRAL antibody
    • Skeletal muscle LIM protein 3 antibody
    • Skeletal muscle LIM-protein 3 antibody
    • SLIM 3 antibody
    • SLIM-3 antibody
    • SLIM3 antibody
    see all

Images

  • Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    All lanes : Anti-FHL2 antibody [EPR17860-23] (ab202586) at 1/1000 dilution

    Lane 1 : Wild-type HeLa lysate
    Lane 2 : FHL2 knockout HeLa lysate
    Lane 3 : K562 lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 32 kDa



    Lanes 1-3: Merged signal (red and green). Green - ab202586 observed at 32 kDa. Red - loading control ab7291 observed at 50 kDa.

     ab202586 Recombinant Anti-FHL2 antibody [EPR17860-23] was shown to specifically react with FHL2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265475 (knockout cell lysate ab257441) was used. Wild-type and FHL2 knockout samples were subjected to SDS-PAGE. ab202586 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 1000 dilution and 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-FHL2 antibody [EPR17860-23] (ab202586)
    Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)

    ab202586 staining FHL2 in wild-type U2OS cells (top panel) and FHL2 knockout U2OS cells (bottom panel). The cells were fixed with 4% PFA (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab202586 at 1/100 dilution and ab7291 (Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo color red).  Nuclear DNA was labelled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)

    ab202586 staining FHL2 in wild-type U2OS cells (top panel) and FHL2 knockout U2OS cells (bottom panel). The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab202586 at 1/100 dilution and ab7291 (Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo color red).  Nuclear DNA was labelled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    All lanes : Anti-FHL2 antibody [EPR17860-23] (ab202586) at 1/2000 dilution

    Lane 1 : SW480 (Human colorectal adenocarcinoma cell line) whole cell lysate
    Lane 2 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate
    Lane 3 : HT1080 (Human fibrosarcoma cells) whole cell lysate
    Lane 4 : HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

    Predicted band size: 32 kDa
    Observed band size: 32 kDa


    Exposure time: 3 minutes


    5% NFDM/TBST: Blocking and diluting buffer.

  • Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Anti-FHL2 antibody [EPR17860-23] (ab202586) at 1/10000 dilution + Human fetal heart lysate at 10 µg

    Secondary
    Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

    Predicted band size: 32 kDa
    Observed band size: 32 kDa


    Exposure time: 30 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Western blot - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    All lanes : Anti-FHL2 antibody [EPR17860-23] (ab202586) at 1/10000 dilution

    Lane 1 : Mouse heart lysate
    Lane 2 : Mouse kidney lysate
    Lane 3 : Rat heart lysate
    Lane 4 : Rat kidney lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

    Predicted band size: 32 kDa
    Observed band size: 32 kDa


    Exposure time: 30 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A-673 (Human muscle Ewing's Sarcoma cell line) cells labeling FHL2 with ab202586 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution (green). Confocal image showing cytoplasmic and weakly nuclear staining on A-673 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/500 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab202586 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/500 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution.

  • Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Immunocytochemistry/ Immunofluorescence - Anti-FHL2 antibody [EPR17860-23] (ab202586)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embyro fibroblast cells) cells labeling FHL2 with ab202586 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution (green). Confocal image showing cytoplasmic and weakly nuclear staining on A-673 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/500 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab202586 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/500 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution.

  • Immunoprecipitation - Anti-FHL2 antibody [EPR17860-23] (ab202586)
    Immunoprecipitation - Anti-FHL2 antibody [EPR17860-23] (ab202586)

    FHL2 was immunoprecipitated from 1mg of SW480 (Human colorectal adenocarcinoma cell line) whole cell lysate with ab202586 at 1/20 dilution. Western blot was performed from the immunoprecipitate using ab202586 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: SW480 whole cell lysate 10 µg (Input). Lane 2: ab202586 IP in SW480 whole cell lysate. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab202586 in SW480 whole cell lysate.

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

    Exposure time: 5 seconds.

References

Publishing research using ab202586? Please let us know so that we can cite the reference in this datasheet.

ab202586 has not yet been referenced specifically in any publications.

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