Thymus chromatin was extensively modified with ethyl acetimidate, substituting up to 90% of the lysyl residues of the histones while retaining the positive charge of the basic amino acid. Physiochemical and immunochemical characterization of this derivative chromatin indicates a high degree of retention of the native structure of the nucleoprotein even after extensive modification. The alterations which are detected are most simply interpreted as resulting from a weakening of the interactions of histone H1 with DNA in the modified chromatin. The near-native character of amidinated chromatin contrasts with the more extensive structural alterations observed in acetylated chromatin. Our data demonstrate the suitability of this reagent for mapping available lysyl residues in this and other nucleoproteins and suggest that the related bisimido esters may be reagents of choice for cross-linking of chromatin histones.