Anti-Lysine Acetylated Antibody was prepared from monospecific antiserum by immunoaffinity chromatography using acetylated lysine peptide coupled to agarose. Assay by immunoelectrophoresis resulted in a single precipitin arc against anti-Rabbit IgG. The antibody reacts specifically with acetylated lysine residues.
Formulierung:
Liquid (sterile filtered)
Application Verdünnung:
ELISA: 1:1,000-1:2,500, IP: 10 µg/mg protein sample, WB: 1:500-1:1,000
Anwendungsbeschreibung:
Application Notes: Anti-Lysine Acetylated Antibody is suitable for use in ELISA, western blotting, immunofluorescence microscopy, and immunoprecipitation assays. Although not tested, this antibody is likely functional in RIA, flow cytometry, and immunohistochemistry
Acetylation does not crosstalk with IR-induced pSer824 and does not affect the co-repressor activity of KAP1.(A) Control and SIRT1 depleted HEK293T cells were harvested at indicated time points following 10Gy-IR. IR-induced KAP1 pSer824 was examined by phosphorylation specific antibody. (B) Cells transfected with FLAG-tagged WT-KAP1 or 4KR mutant were harvested at indicated time points following 4Gy-IR. Exogenous KAP1 was immunoprecipitated and pSer824 was examined by phosphorylation specific antibody. (C) Cyclin D3 luciferase construct was co-transfected with combinations of HA-tagged E2F1, FLAG-tagged KAP1, and 4KR mutant. Luciferase activity was measured 48 hours post transfection. (D) Cells transfected with FLAG-tagged WT-KAP1 or KAP1 phospho mutants (S824A and S824D) were harvested at indicated time points following 10Gy-IR. Recombinant KAP1 was purified by immunoprecipitation and the total acetylation level was assessed by immunoblotting.
RAD54 acetylation is important for BRD9 recognition and HR activity. a-c RAD54 is acetylated by GCN5/PCAF and deacetylated by HDAC 6/HDAC11 following induction of DNA damage. a 293T cells were transfected with control or RAD54-HA plasmid. Twenty-four hours after transfection, cells were exposed to the 10-Gy IR and harvested at the indicated time points. Immunoprecipitation with anti-HA beads was performed. Blots were probed with the indicated antibodies. b 293T cells were transfected with the indicated plasmids. Twenty-four hours after transfection, cells were exposed to 10-Gy IR, and lysates were collected after 8 h. Immunoprecipitation with anti-HA beads was performed. Blots were probed with the indicated antibodies. c 293T cells were transfected with the indicated plasmids, treated as indicated, and subjected to immunoprecipitation as outlined in b. Blots were probed with the indicated antibodies. d, e RAD54 K515 acetylation is important for RAD51-RAD54 complex formation. d 293T cells were transfected with the indicated plasmids, treated as indicated, and subjected to immunoprecipitation as outlined in b. Blots were probed with the indicated antibodies. e 293T cells were transfected with the indicated plasmids, exposed to either no IR or 10-Gy IR as indicated, and subjected to immunoprecipitation as outlined in b. Blots were probed with the indicated antibodies. f-h RAD54 K515 acetylation is essential for HR activity. f, g U2OS cells were transfected with the indicated plasmids and exposed to 2-Gy IR. Cells were fixed after 8 h and stained for the indicated proteins. Representative immunofluorescence images of RAD51 (green) and RAD54 (red) are shown in f. Quantification of the indicated foci is shown in g. Representative data (mean SEM) are shown from n = 50 cells examined over three independent experiments. **p < 0.01, ***p < 0.001 by two-sided unpaired t test. NS not significant. Scale bar, 10 µm. h Survival curves of U2OS cells expressing the indicated constructs and exposed to the indicated doses of PARPi or cisplatin. Representative data (mean SEM) are shown from n = 3 biologically independent samples. *p < 0.05, ***p < 0.001 by two-sided unpaired t test.
RAD54 acetylation is important for BRD9 recognition and HR activity. a-c RAD54 is acetylated by GCN5/PCAF and deacetylated by HDAC 6/HDAC11 following induction of DNA damage. a 293T cells were transfected with control or RAD54-HA plasmid. Twenty-four hours after transfection, cells were exposed to the 10-Gy IR and harvested at the indicated time points. Immunoprecipitation with anti-HA beads was performed. Blots were probed with the indicated antibodies. b 293T cells were transfected with the indicated plasmids. Twenty-four hours after transfection, cells were exposed to 10-Gy IR, and lysates were collected after 8 h. Immunoprecipitation with anti-HA beads was performed. Blots were probed with the indicated antibodies. c 293T cells were transfected with the indicated plasmids, treated as indicated, and subjected to immunop
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