Anti-eIF4A2 antibody [EPR27347-74]

16 product images

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  Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

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

  This antibody does not cross-react with human EIF4A1 and human EIF4A3.

  This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

  In Western blot, anti-His antibody (Anti-6X His tag® antibody [EPR20547] - ChIP Grade ab213204) staining at 1/5000 dilution.

  All lanes: Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728) at 1/1000 dilution

  Lane 1: His-tagged human EIF4A1 recombinant protein at 10 ng

  Lane 2: His-tagged human EIF4A2 recombinant protein at 10 ng

  Lane 3: His-tagged human EIF4A3 recombinant protein at 10 ng

  Secondary

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

  Developed using the ECL technique.

  Observed band size: 48 kDa

  Exposure time: 48s

• 

  Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Western blot: Anti-EIF4A2 antibody [EPR27347-74] (ab307728) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (Anti-Calnexin antibody [CANX/1543] ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab307728 was shown to bind specifically to EIF4A2. A band was observed at 48 kDa in wild-type A549 cell lysates with no signal observed at this size in EIF4A2 knockout cell line. To generate this image, wild-type and EIF4A2 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

  All lanes: Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728) at 1/1000 dilution

  Lane 1: Wild-type A549 cell lysate at 20 µg

  Lane 2: EIF4A2 knockout A549 cell lysate at 20 µg

  Lane 3: MCF7 cell lysate at 20 µg

  Lane 4: MCF7 Membrane Prep cell lysate at 20 µg

  Secondary

  Lanes 1 - 4: Goat anti-Rabbit IgG H&L 800CW at 1/20000 dilution

  Lanes 1 - 4: Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

  Performed under reducing conditions.

  Observed band size: 48 kDa

• 

  Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST
  The identity of the lower MW band at approximately 20 kDa is unknown.
  Exposure time: 15 seconds

  All lanes: Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728) at 1/1000 dilution

  Lane 1: C6 (rat glial tumor glial cell) whole cell lysate at 20 µg

  Lane 2: PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg

  Lane 3: RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

  Lane 4: C2C12 (mouse myoblast) whole cell lysate at 20 µg

  Secondary

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

  Observed band size: 48 kDa

  Exposure time: 15s

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunohistochemical analysis of paraffin-embedded Rat testis tissue labeling eIF4A2 with ab307728 at 1/10000 (0.05 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining on rat testis. The section was incubated with ab307728 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background. Counterstained with Hematoxylin.
  Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
  Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins

• 

  Immunocytochemistry/ Immunofluorescence - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized C6 (rat glial tumor glial cell) cells labelling eIF4A2 with ab307728 at 1/50 (9.92 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green). Confocal image showing cytoplasmic and very weak nuclear staining in C6 cell line.Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• 

  Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST
  
  Exposure time: 10 seconds

  All lanes: Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728) at 1/1000 dilution

  Lane 1: HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 2: MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 3: 293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

  Lane 4: Raji (human Burkitts lymphoma B lymphocyte) whole cell lysate at 20 µg

  Lane 5: Human testis tissue lysate at 20 µg

  Secondary

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

  Observed band size: 48 kDa

  Exposure time: 10s

• 

  Immunoprecipitation - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  eIF4A2 was immunoprecipitated from 0.35 mg HeLa whole cell lysate with ab307728 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307728 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
  Lane 1: HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate
  Lane 2: ab307728 IP in HeLa whole cell lysate
  Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab307728 in HeLa whole cell lysate
  Blocking and dilution buffer and concentration: 5% NFDM/TBST.
  Exposure time: 1 seconds

  All lanes: Immunoprecipitation - Anti-eIF4A2 antibody [EPR27347-74] (ab307728) at 1/30 dilution

  Secondary

  All lanes: Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) at 1/5000 dilution

  Exposure time: 1s

• 

  Immunocytochemistry/ Immunofluorescence - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells labelling eIF4A2 with ab307728 at 1/50 (9.92 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green). Confocal image showing cytoplasmic and weak nuclear staining in HeLa cell line.Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

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  Flow Cytometry (Intracellular) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized C6 (rat glial tumor glial cell) cells labelling eIF4A2 with ab307728 at 1/500 dilution (0.1ug) (Red) (Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.

• 

  Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST
  Low expression: liver(PMID:8521730).
  
  In Western blot, anti- H3 antibody (Anti-Histone H3 antibody [EPR16987] - Nuclear Marker and ChIP Grade ab176842) loading control staining at 1/100000 dilution.
  Exposure time: 6 seconds

  All lanes: Western blot - Anti-eIF4A2 antibody [EPR27347-74] (ab307728) at 1/1000 dilution

  Lane 1: Human brain tissue lysate at 20 µg

  Lane 2: Human heart tissue lysate at 20 µg

  Lane 3: Human liver tissue lysate at 20 µg

  Secondary

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

  Observed band size: 48 kDa

  Exposure time: 6s

• 

  Immunocytochemistry/ Immunofluorescence - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized Raw 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cells labelling eIF4A2 with ab307728 at 1/50 (9.92 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green). Confocal image showing cytoplasmic and weak nuclear staining in Raw 264.7 cell line.Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• 

  Flow Cytometry (Intracellular) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cells labelling eIF4A2 with ab307728 at 1/500 dilution (0.1ug) (Red) (Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labeling eIF4A2 with ab307728 at 1/10000 (0.05 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining on mouse testis. The section was incubated with ab307728 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background. Counterstained with Hematoxylin.
  Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
  Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunohistochemical analysis of paraffin-embedded Human breast carcino tissue labeling eIF4A2 with ab307728 at 1/10000 (0.05 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining on human breast carcinoma (PMID: 21219636). The section was incubated with ab307728 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background. Counterstained with Hematoxylin.
  Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
  Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins

• 

  Flow Cytometry (Intracellular) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labelling eIF4A2 with ab307728 at 1/500 dilution (0.1ug) (Red) (Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eIF4A2 antibody [EPR27347-74] (ab307728)

  Immunohistochemical analysis of paraffin-embedded Human testis tissue labeling eIF4A2 with ab307728 at 1/10000 (0.05 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining on human testis. The section was incubated with ab307728 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background. Counterstained with Hematoxylin.
  Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
  Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins