There are several factors one should consider in order to get the best ChIP results in their experiment.
- Use an appropriate control, like an isotype control or non-related antibody, to assess antibody specificity.
- Ensure that the antibody is validatedfor ChIP applications, or use one validated for immunoprecipitation (IP).
- Optimize the antibody-to-chromatin ratiofor efficient immunoprecipitation.
- Use polyclonal antibodies or epitope tagsto reduce false negatives.
- Use the right chromatin preparation methodbased on the target proteins, such as xChIP for weak DNA binding.
- Confirm chromatin fragmentationusing agarose gel electrophoresis after DNA purification.
- Compare protein enrichment of target sequencesto unrelated regions to validate results.
- Preserve post-translational modificationswith specific inhibitors to stop degradation of the modifications on proteins.
- Verify ChIP DNA quality(at a minimum of 5-fold enrichment) through PCR or qPCR, comparing them to controls.