AbFlex^® Histone H3K9me0 antibody (rAb)

Application Notes

Applications Validated by Active Motif:
ICC/IF: 1 - 2 µg/ml
WB*: 0.1 - 1 µg/ml
Bead-based ELISA: 0.5 - 10 µg/ml

*Note: many chromatin-bound proteins are not soluble in a low salt nuclear extract and fractionate to the pellet. Therefore, we recommend a High Salt / Sonication Protocol when preparing nuclear extracts for Western Blot.

Immunogen

This antibody was raised against a synthetic branched peptide corresponding to amino acids surrounding Lys9 of human Histone H3.

Buffer

Purified IgG in 140 mM Hepes, pH 7.5, 70 mM NaCl, 32 mM NaOAc, 30% glycerol and 0.035% sodium azide. Sodium azide is highly toxic.

Background

AbFlex^® antibodies are recombinant antibodies (rAbs) that have been generated using defined DNA sequences to produce highly specific, reproducible antibodies. Each AbFlex antibody contains a 6xHis Tag, a Biotinylation Tag for enzymatic biotin conjugation using the biotin ligase, BirA, and a sortase recognition motif (LPXTG) to attach a variety of labels directly to the antibody including fluorophores, enzymatic substrates (HRP, AP), peptides, drugs as well as solid supports.

AbFlex^® Histone H3K9me0 antibody was expressed as full-length IgG with mouse immunoglobulin heavy and light chains (IgG2a isotype) in mammalian 293 cells.

Histone H3 is one of the core components of the nucleosome, the basic building block of chromatin. Histones are subject to a variety of chemical modifications, including post-translational modifications of the histone proteins and the methylation of cytosine residues in the DNA. Reported histone modifications include acetylation, methylation, phosphorylation, ubiquitylation, glycosylation, ADP-ribosylation, carbonylation and SUMOylation; these modifications play a major role in regulating gene expression.

Methylation of lysines occur as four states: unmethylated (me0), monomethyl (me1), dimethyl (me2) and trimethyl (me3). The me0 state of lysine is recognized as biologically relevant and a number of proteins containing PhD fingers, ADD and WD40 domains are known to associate with unmodified lysines.

Positive Control

Active Motif's HeLa acid extract (Catalog No. 36200) can be used as a positive control for Histone H3K9me0 antibody.

Chromatin IP Assays

This antibody has been validated for use in ChIP and/or ChIP-Seq, and can be used with Active Motif's ChIP-IT^® High Sensitivity Kit or our magnetic bead-based ChIP-IT^® Express Kits. For an overview of all of our ChIP products, please visit our Chromatin IP Page to learn about ChIP products designed for use with AbFlex^® Histone H3K9me0 antibody (rAb).

Storage

Some products may be shipped at room temperature. This will not affect their stability or performance. Avoid repeated freeze/thaw cycles by aliquoting items into single-use fractions for storage at -20°C for up to 2 years. Keep all reagents on ice when not in storage.

Guarantee

This product is guaranteed for 12 months from date of receipt.

This product is for research use only and is not for use in diagnostic procedures.

Application Key

• ChIP = Chromatin Immunoprecipitation;
• ChIP = ChIP Sequencing;
• CUT&RUN = Cleavage Under Targets and Release Using Nuclease;
• CUT&Tag = Cleavage Under Targets and Tagmentation;
• DB = Dot Blot;
• ELISA = Enzyme-linked Immunosorbent Assay;
• EMSA = Electrophoretic Mobility Shift Assay
• FC = Flow Cytometry;
• ICC = Immunocytochemistry;
• IF = Immunofluorescence;
• IHC = Immunohistochemistry;
• IP = Immunoprecipitation;
• MeDIP = Methyl-DNA Immunoprecipitation;
• NEU = Neutralizing;
• WB = Western Blotting