Anti-CREB antibody [LB9]

9 product images

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  Western blot - Anti-CREB antibody [LB9] (ab178322)

  Lane 1: Wild-type HAP1 cell lysate (20 μg)
  Lane 2: CREB knockout HAP1 cell lysate (20 μg)
  Lane 3: HeLa cell lysate (20 μg)
  Lane 4: A431 cell lysate (20 μg)
  Lanes 1 - 4: Merged signal (red and green). Green - ab178322 observed at 44 kDa. Red - loading control, ab18058, observed at 124 kDa.
  
  ab178322 was shown to recognize CREB when CREB knockout samples were used, along with additional cross-reactive bands. Wild-type and CREB knockout samples were subjected to SDS-PAGE. ab178322 and ab18058 (loading control to Vinculin) were diluted at 1/250 and 1/10,000 dilution respectively and incubated overnight at 4C. Blots were developed with IRDye® 800CW Goat anti-Rabbit IgG (H + L) and IRDye® 680 Goat anti-Mouse IgG (H + L) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

  All lanes: Western blot - Anti-CREB antibody [LB9] (ab178322)

  Predicted band size: 36 kDa

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  Immunoprecipitation - Anti-CREB antibody [LB9] (ab178322)

  Immunoprecipitation analysis of CREB was performed on untreated 293T cells. The antigen:antbody complex was formed by binding 500μg whole cell lysate with 2μg ab178322 and captured on 50μl Protein A/G Plus Agarose.

  Lane 1: 293T lysate

  Lane 2: IP of 293T lysate

  WB detection used ab178322 at 1/1000 followed with a goat anti-mouse-HRP secondary antibody at 1/20,000.

  All lanes: Immunoprecipitation - Anti-CREB antibody [LB9] (ab178322)

  Developed using the ECL technique.

  Predicted band size: 36 kDa

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  Western blot - Anti-CREB antibody [LB9] (ab178322)

  All lanes: Western blot - Anti-CREB antibody [LB9] (ab178322) at 1/500 dilution

  Lane 1: Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 30 µg

  Lane 2: HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 30 µg

  Lane 3: K-562 (Human chronic myelogenous leukemia lymphoblast cell line ) whole cell lysate at 30 µg

  Lane 4: Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 30 µg

  Lane 5: NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 30 µg

  Lane 6: C2C12 whole cell lysate at 30 µg

  Lane 7: U-2 OS (Human bone osteosarcoma epithelial cell line) whole cell lysate at 30 µg

  Lane 8: A549 (Human lung carcinoma cell line) whole cell lysate at 30 µg

  Lane 9: MDA-MB-231 (Human breast adenocarcinoma cell line) whole cell lysate at 30 µg

  Lane 10: T98G whole cell lysate at 30 µg

  Predicted band size: 36 kDa

  Observed band size: 43 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CREB antibody [LB9] (ab178322)

  Immunohistochemical analysis of deparaffinized Human tonsil tissue labeling CREB with ab178322 at 1/100 dilution followed by DAB staining.

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  Western blot - Anti-CREB antibody [LB9] (ab178322)

  All lanes: Western blot - Anti-CREB antibody [LB9] (ab178322) at 1/500 dilution

  Lane 1: HeLa whole cell lysate at 25 µg

  Lane 2: 293T whole cell lysate at 25 µg

  Lane 3: A431 whole cell lysate at 25 µg

  Lane 4: A549 whole cell lysate at 25 µg

  Lane 5: MEF whole cell lysate at 25 µg

  Lane 6: MCF7 whole cell lysate at 25 µg

  Lane 7: U2OS whole cell lysate at 25 µg

  Lane 8: K562 whole cell lysate at 25 µg

  Lane 9: COS7 whole cell lysate at 25 µg

  Lane 10: 3T3L1 whole cell lysate at 25 µg

  Lane 11: C2C12 whole cell lysate at 25 µg

  Lane 12: NRK whole cell lysate at 25 µg

  Secondary

  All lanes: goat anti-mouse-HRP at 1/20000 dilution

  Developed using the ECL technique.

  Predicted band size: 36 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CREB antibody [LB9] (ab178322)

  Immunohistochemical analysis of deparaffinized Human bladder cancer (left) and rectal cancer (right) tissues labeling CREB with ab178322 at 1/200 dilution followed by DAB staining.

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  Immunocytochemistry/ Immunofluorescence - Anti-CREB antibody [LB9] (ab178322)

  Immunofluorescent analysis of untreated HeLa cells (formalin-fixed, 0.1% Triton X-100 permeabilized) labeling CREB with ab178322 at 1/400 dilution (green) followed with DyLight 488 goat anti-mouse IgG secondary antibody at 1/400 dilution. Nuclei (blue) were stained with Hoechst 33342 dye.

• 

  Immunocytochemistry/ Immunofluorescence - Anti-CREB antibody [LB9] (ab178322)

  Immunofluorescent analysis of untreated HeLa cells (formalin-fixed, 0.1% Triton X-100 permeabilized) labeling CREB with ab178322 at 1/500 dilution (green) followed with DyLight 488 goat anti-mouse IgG secondary antibody at 1/400 dilution. Nuclei (blue) were stained with Hoechst 33342 dye.

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  Western blot - Anti-CREB antibody [LB9] (ab178322)

  Western blot: Anti-CREB1 antibody [LB9] (ab178322) staining at 1/500 dilution, shown in green; Rabbit anti-ACTN4 [EPR2533(2)] (Anti-alpha Actinin 4 antibody [EPR2533(2)] ab108198) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab178322 was shown to bind specifically to CREB1. A band was observed at 40 kDa in wild-type A549 cell lysates with no signal observed at this size in CREB1 knockout cell line. To generate this image, wild-type and CREB1 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 3 % 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-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution.

  All lanes: Western blot - Anti-CREB antibody [LB9] (ab178322) at 1/500 dilution

  Lane 1: Wild-type HCT 116 cell lysate at 20 µg

  Lane 2: CREB1 knockout HCT 116 cell lysate at 20 µg

  Lane 3: Wild-type A549 ab288558 cell lysate at 20 µg

  Lane 4: CREB1 knockout A549 Human CREB1 knockout A549 cell line ab286720 cell lysate at 20 µg

  Secondary

  All lanes: Goat anti-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution

  Performed under reducing conditions.

  Predicted band size: 36 kDa

  Observed band size: 40 kDa