Anti-acetyl Lysine antibody - ChIP Grade

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-acetyl Lysine antibody - ChIP Grade (AB21623)

Immunofluorescent staining of Human melanoma cells, using Rabbit polyclonal to acetyl Lysine (ab21623) at 1 : 100 dilution.

1 : Untreated cells

2 : TSA treated cells

• IP

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Immunoprecipitation - Anti-acetyl Lysine antibody - ChIP Grade (AB21623)

p53 acetylation upon Doxorubicin treatment in human melanoma cells (MMRU cells). MMRU cells were treated with 0.5 ug/ml Dox for various times and lyzed for whole protein. Immunoprecipitation was performed with ab21623. Western blot was performed to detect the immunoprecipitated p53 with anti-p53 antibody.

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Immunoprecipitation - Anti-acetyl Lysine antibody - ChIP Grade (ab21623)

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• ChIP

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ChIP - Anti-acetyl Lysine antibody - ChIP Grade (AB21623)

Chromatin was prepared from Hela cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The ChIP was performed with 25μg of chromatin, 2μg of ab21623 (blue), and 20μl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region.

• WB

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Western blot - Anti-acetyl Lysine antibody - ChIP Grade (AB21623)

Lane 1 = Extract of Mcf7 cells incubated with vehicle 20 ug. Lane 2 = Extract of Mcf7 cells incubated with trichostatin A 20 ug. Lane 3 = Extract of Mcf7 cells incubated with EX527 20 ug. Lane 4 = Extract of Mcf7 cells incubated with nicotinamide 20 ug. Lane 5 = Extract of Mcf7 cells incubated with camptothecin 20 ug. Lane 6 = Extract of Mcf7 cells incubated with camptothecin and trichostatin A 20 ug. Lane 7 = Extract of Mcf7 cells incubated with camptothecin and EX527 20 ug. Lane 8 = Extract of Mcf7 cells incubated with camptothecin and nicotinamide 20 ug. Lane 9 = Extract of 293T cells incubated with vehicle 20 ug. Lane 10 = Extract of 293T cells incubated with camptothecin 20 ug. Lane 11 = Extract of 293T cells incubated with camptothecin and trichostatin A 20 ug. Lane 12 = Extract of 293T cells incubated with camptothecin and EX527 20 ug. Lane 13 = Extract of 293T cells incubated with camptothecin and nicotinamide 20 ug
SDS PAGE performed under reducing conditions (100mM DTT Sample heated at 50°C). Primary : Lanes 1-13 : Rabbit anti acetyl Lysine antibody (ab21623) at 1/500 dilution. Secondary : Lanes 1-13 : Goat anti rabbit IgG(H&L)-IR680 at 1 : 10,000 (in green). Developed : Oddysey. Blocking : in 5% Milk + PBS for 3 hours at RT. Primary antibody : in 5% BSA + 50mM Tris pH 7.5 + 150 mM NaCl + 0.05% Tween-20. Secondary antibody : in 5% Milk + PBS + 0.1% Tween-20 + 0.01%SDS for 2 hour at RT. Predicted band size : multiple. Observed band size : multiple

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Western blot - Anti-acetyl Lysine antibody - ChIP Grade (ab21623)

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• WB

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Western blot - Anti-acetyl Lysine antibody - ChIP Grade (AB21623)

Primary : All Lanes : Anti acetyl Lysine antibody (ab21623) at 1 : 1000. Lane 1 : Marker. Lane 2 : HeLa cells vehicle-treated (ab139414). Lane 3 : HeLa cells, trichostatin A-treated (ab139414). Lysates at 20 ug/lane . Secondary : All Lanes : Goat anti-Rabbit IgG 1 : 10000. Performed under reducing conditions. Blocking buffer : 5% milk in PBS. Observed band sizes : 11 kDa 15kDa 45kDa 50 kDa.

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Western blot - Anti-acetyl Lysine antibody - ChIP Grade (ab21623)

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• WB

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Western blot - Anti-acetyl Lysine antibody - ChIP Grade (AB21623)

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Western blot - Anti-acetyl Lysine antibody - ChIP Grade (ab21623) at 0.5 µg/mL

Lane 1:

Untreated Human melanoma cell lysate at 75 µg

Lane 2:

TSA-treated Human melanoma cell lysate at 75 µg

Secondary

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Goat anti-rabbit IgG HRP at 0.25 µg/mL

Observed band size: 14-Dec kDa,16-18 kDa

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