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Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures.
Successful immunoblotting of histones must overcome an important consequences of histone composition, namely, their relatively low molecular weight. The molecular weight of histones effects both resolution of histone proteins in SDS-PAGE and their efficient transfer to nitrocellulose membrane. Good results are achieved by using high percent agarose gels and nitrocellulose with a pore size of 0.2 um to ensure optimal capture of histone proteins. We also suggest using high quality BSA in the blocking solutions rather than conventional dried milk.
Gel Electrophoresis of Proteins: A Practical Approach. Third Edition. Ed. B.D. Hames. Oxford University Press. Oxford, UK. 1998
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