Double labeling of chromatin proteins,in vivo and in vitro,and their two-dimensional electrophoretic resolution

A modification of the two-dimensional electrophoretic method that involves nonequilibrium pH gradients has been adapted for high resolution of chromatin proteins from sea urchin embryos. A simple method of labeling the protein, in vitro, by reductive methylation with boro³H]hydride to a specific activity of 100,000 cpm/μg of protein is detailed. Chromatin protein may be labeled, in vivo with ¹⁴C-amino acids, and newly synthesized (³H and ¹⁴C-labeled) and preexistent proteins (only ³H labeled) may be distinguished. The method reveals that sea urchin embryo chromatin contains over 200 proteins.