Acetyl-Lysine Affinity Beads - AAC02

Acetyl-Lysine Affinity Beads

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AAC02-Beads

Cytoskeleton offers two types of Acetyl-Lysine Affinity Beads, Cat # AAC02-Beads and Cat # AAC03-Beads. Both reagents are comprised of mouse anti-acetyl-lysine antibodies that are covalently linked to protein G beads and both enrich a broad range of acetylated proteins. AAC02-Beads and AAC03-Beads can be combined to give a more extensive acetylated protein enrichment profile (Cat # AAC04-Beads is a 1:1 combination of AAC02-Beads:AAC03-Beads).

AAC02-Beads has an overlapping but unique specificity profile when compared to AAC03-Beads and may outperform AAC03-Beads when examining a specific target protein, see Detailed methods and Validated Applications for examples. When examining the acetylation state of a new protein of interest (POI), it is recommended to try AAC03-Beads and AAC02-Beads separately to determine which reagent is optimal for your POI.

Each lot of affinity-bead is quality controlled to provide high batch to batch consistency, see COA documents.

Validated Applications

Application 1: Detection of Acetylation Changes in a Target POI

AAC02-Beads have been shown to be a superior reagent for the immunoprecipitation of acetylated PDHE1 (Fig. 1). In the presence of hydrogen peroxide AAC02 Bead IPs show a 2.6 fold de-acetylation of PDHE1 while AAC03 Bead shows a 1.5 fold de-acetylation and 8-9 fold weaker signal. The specificity of the beads for acetylated PDHE1 is shown by the lack of signal from mouse IgG control beads (Fig. 1). A potential mechanism for deacetylation in response to hydrogen peroxide treatment is the upregulation of the deacetylase SIRT3.

Figure 1 Legend: AAC02-Beads & AAC03-Beads (50μl bead slurry) were used to IP acetylated proteins from A431 cell lysates either treated (+) or untreated (-) with hy-drogen peroxide (100 μM) for 2 hours. Each IP used 1 mg of lysate. Western blot analysis using anti PDHE1 antibody was performed and signals were quantitated using LiCor Empiria software (Table). Mouse IgG control beads (Cat #CIG02) and AAC04-Beads (25 μl AAC02-Beads & 25 μl AAC03-Beads) were also used in IP assays (1 mg lysate per IP). Input lanes repre-sent 2% of IP input lysate (20 μg) from treated (+) or untreated (-) lysate. Input signal represents total PDHE1.

 

 

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Application 2: Immunoprecipitation of Total Acetyl-Lysine Profile

AAC02-Beads show a robust total acetyl IP profile with an overlapping but unique specificity profile to AAC03-Beads.  AAC02-Beads can be used alone or in combination with AAC03-Beads to study the acetylome.

Figure 2 Legend: AAC02-Beads, AAC03-Beads, and AAC04-Beads (50µl bead slurry) were used to IP acetylated proteins from Cos-7 cells either treated (+) or untreated (-) with deacetylase inhibitors [TSA (1µM) and nicotinamide (1mM)] for 6 hours. The total profile of enriched acetylated proteins were eluted and analyzed by western blot with an AAC03-HRP antibody (1:3000).  Mouse IgG beads are used as a control for non-specific binding (Cat # CIG02). Each IP assay utilized 1 mg of Cos-7 lysate.

 

 

aac04_fig1_2

Amount:
Each package contains enough acetyl-lysine affinity beads for 40 reactions.

 

For more information contact: signalseeker@cytoskeleton.com

Associated Products:

Signal-Seeker™ SUMOylation 2/3 Detection Kit (Cat. # BK162)

Signal-Seeker™ Ubiquitination Detection Kit (Cat. # BK161)

Signal-Seeker™ Acetyl-Lysine Detection Kit (Cat. # BK163)

Signal-Seeker™: BlastR™ Rapid Lysate Prep Kit (Cat. # BLR01)

Signal-Seeker™: PTMtrue™ Aceetyl lysine Antibody (Cat.# AAC03)

For product Datasheets and MSDSs please click on the PDF links below.   For additional information, click on the FAQs tab above or contact our Technical Support department at tservice@cytoskeleton.com

AuthorTitleJournalYearArticle Link
He, Ying et al.PPARγ Acetylation Orchestrates Adipose Plasticity and Metabolic RhythmsAdvanced Science2023ISSN 2198--3844
Oldfield C. et. al.Muscle-specific sirtuin 3 overexpression does not attenuate the pathological effects of high-fat/high-sucrose feeding but does enhance cardiac SERCA2a activityPhysiol Rep.2021ISSN 3440-5591
Kumar, Manish et al.Inhibition of histone acetyltransferase function radiosensitizes CREBBP/EP300 mutants via repression of homologous recombination, potentially targeting a gain of functionNature Communications2021ISSN 2041-1723
Horita, Henrick et al.Utilizing a comprehensive immunoprecipitation enrichment system to identify an endogenous post-translational modification profile for target proteinsJournal of Visualized Experiments2018ISSN 1940-087X
Horita, Henrick et al.Utilizing optimized tools to investigate PTM crosstalk: Identifying potential PTM crosstalk of acetylated mitochondrial proteinsProteomes2018ISSN 2227-7382

If you have any questions concerning this product, please contact our Technical Service department at tservice@cytoskeleton.com