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Methods Mol Biol. 2009;567:27-43. doi: 10.1007/978-1-60327-414-2_2.

Characterization and quality control of antibodies used in ChIP assays.

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1
Diagenode sa, Sart-Tilman, Li├Ęge, Belgium.

Abstract

We present here the very robust characterization and quality control (QC) process that we have established for our polyclonal antibodies, which are mainly directed against targets relevant to the epigenetics field such as modified histones, modifying enzymes, and chromatin-interacting proteins. The final purpose of the characterization and QC is to label antibodies as chromatin immunoprecipitation (ChIP) grade. Indeed, the ChIP method is extensively used in epigenetics to study gene regulation and relies on the use of antibodies to select the protein of interest and then precipitate and identify the DNA associated to it. We have optimized in-house all protocols and reagents needed from the first to the last step of antibody characterization. First, following immunizations, the rabbit crude serum is tested for immune response. Whether or not the antibody is specific is determined in further characterizations. Then, only specific antibodies are tested in ChIP using an optimized method which is ideal for antibody screening. Once QC is established for one antibody, it is used to similarly characterize each antibody batch in order to supply researchers in a reproducible manner with validated antibodies. All in all, this demonstrates that we develop epigenetics research tools based on everyday's researcher's needs by providing batch-specific fully characterized ChIP-grade antibodies.

PMID:
19588083
DOI:
10.1007/978-1-60327-414-2_2
[Indexed for MEDLINE]
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