Acetyl-Lysine Antibody (1C6)

Western Blot: Acetyl-Lysine Antibody (1C6) [NB100-74339]

Western Blot: Acetyl-Lysine Antibody (1C6) [NB100-74339] - Analysis of Acetyl-Lysine was performed by loading various amounts of acetylated BSA or non-acetylated BSA per well and 10ul of PageRuler Prestained Protein Ladder onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% BSA/TBST for at least 1 hour. The membrane was probed with an Acetyl-Lysine monoclonal antibody at a dilution of 1:1000 overnight at 4C on a rocking platform, washed in TBS-0.1%Tween-20, and probed with a goat anti-mouse IgG-HRP secondary antibody at a dilution of 1:20,000 for 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico.

Western Blot: Acetyl-Lysine Antibody (1C6) [NB100-74339]

Western Blot: Acetyl-Lysine Antibody (1C6) [NB100-74339] - Analysis of lysine acetylated proteins from cells left untreated (DMSO only) or cells treated with 0.3uM or 3uM of Trichostatin A (TSA) for 16 hours was performed by loading 50 ug of the indicated whole cell lysates per well and 10ul of PageRuler Prestained Protein Ladder onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% BSA/TBST for at least 1 hour. The membrane was probed with an Acetyl-Lysine monoclonal antibody at a dilution of 1:1000 overnight at 4C on a rocking platform, washed in TBS-0.1%Tween-20, and probed with a goat anti-mouse IgG-HRP secondary antibody at a dilution of 1:20,000 for 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico.

Dot Blot: Acetyl-Lysine Antibody (1C6) [NB100-74339] - Specific detection of lysine acetylated histones using an Acetyl-Lysine monoclonal antibody. Histone peptides containing 384 different modification combinations spotted in duplicate on a peptide array were used according to the manufacturer instructions. A schematic and table with representative results (positive detection in red, negative in blue) are shown. Result interpretation and quantification were performed using the manufacturer software. Chemiluminescent detection was performed.

Immunoprecipitation: Acetyl-Lysine Antibody (1C6) [NB100-74339]

Immunoprecipitation: Acetyl-Lysine Antibody (1C6) [NB100-74339] - Immunoprecipitation of acetylated alpha-Tubulin was performed.3uM or 3uM Trichostatin A (TSA) for 16 hours. Antigen-antibody complexes were formed by incubating 500ug of lysate with 3ug of an Acetyl-Lysine monoclonal antibody overnight on a rocking platform at 4C. The immune complexes were captured on 50ul Protein A/G Agarose at a dilution of 1:1000 overnight rotating at 4C, washed in TBST, and probed with Clean-blot IP Detection Reagent at a dilution of 1:2000 for at least 1 hour.

Immunoprecipitation: Acetyl-Lysine Antibody (1C6) [NB100-74339]

Immunoprecipitation: Acetyl-Lysine Antibody (1C6) [NB100-74339] - Analysis of acetylated Histone H3 (Lys9) was performed using whole cell lysates from cells left untreated (DMSO only) or cells treated with 0.3uM or 3uM Trichostatin A (TSA) for 16 hours. Antigen-antibody complexes were formed by incubating 500ug of the indicated lysate with 3ug of an Acetyl-Lysine mAb overnight on a rocking platform at 4C. The immune complexes were captured on 50ul Protein A/G Agarose, washed extensively, and eluted with 5X Lane Marker Reducing Sample Buffer. Samples were resolved on a 4-20% Tris-HCl polyacrylamide gel, transferred to a PVDF membrane, and blocked with 5% BSA/TBS-0.1%Tween for at least 1 hr. The membrane was probed with an Acetyl-Histone H3 (Lys9) mAb at a dilution of 1:1000 overnight rotating at 4C, washed in TBST, and probed with Clean-blot IP Detection Reagent at a dilution of 1:2000 for at least 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico.

Dot Blot: Acetyl-Lysine Antibody (1C6) [NB100-74339] - Specific detection of lysine acetylated histones using an Acetyl-Lysine monoclonal antibody. Histone peptides containing 384 different modification combinations spotted in duplicate on a peptide array were used according to the manufacturer's instructions (Active Motif). Representative results showing specific detection of acetylated Histone H4 K20 (blue circles, panel A) and H3 K9 (blue circles, panel B). Note that Histone H4 K20me2/K20me3 (red and yellow circles, panel A) and H3 K9me2/K9m3 (red and yellow circles, panel B) are not detected by the Acetyl-Lysine monoclonal antibody.