Nucleosome ELISA (NU-ELISA) is a sensitive and quantitative method to detect global patterns of post-translational modifications in preparations of native, intact nucleosomes. These modifications include methylations, acetylations, and phosphorylations at specific histone amino acid residues, and hence NU-ELISA provides a global proteomic assay of the overall chromatin modification states of specific cell types.
Total time
~4–6 hours per sample preparation and ELISA assay
Steps
1
Isolate cell nuclei from tissue or culture
Extract nuclei from cells using standard nuclear isolation protocols. This step prepares intact nuclei for subsequent chromatin digestion.
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2
Digest chromatin with micrococcal nuclease in situ
Treat isolated nuclei with micrococcal nuclease to selectively cleave linker DNA and release native, intact nucleosomes. Optimize digestion to preserve nucleosome structure.
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3
Perform nucleosome-ELISA to detect modifications
Capture nucleosomes on ELISA plates and detect specific histone post-translational modifications using modification-specific antibodies. Quantify signal to determine global modification patterns.
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