The ability of avian-specific linker histone H5, and the globular domains of H5 (GH5) and H1(0) (GH1(0), to self-associate either free in solution or when bound to DNA was investigated. All three proteins underwent a salt-dependent increase in turbidity that may be indicative of nonspecific interactions. Dithiobis(succinimidyl propionate) cross-linking was used to measure specific contacts for both H5 and GH5 free in solution and bound to DNA. H5 and GH5 each became cross-linked in solution, with GH5 displaying divalent polymerization interactions, which suggests that two specific surfaces were involved in the assembly process. For GH5-DNA complexes, cross-linking appeared to be largely the consequence of aggregation, but under low concentrations of DSP, cross-linking GH5 was observed to assemble preferentially onto DNA before oligomerizing to form massive aggregates. Both linear and supercoiled DNA facilitated GH5 interactions compared to assembly in solution; differences in the distribution of cross-linked polymer sizes indicates that assembly is dependent on both the presence of DNA and the morphology of the DNA. Finally, on the basis of a technique referred to as quantitative proteolysis, GH5 assembly on DNA appears to involve specific protein-protein contacts involving the C terminus of one partner. Overall, the cumulative results reported here support the premise that linker histones assemble specifically both in solution and on DNA.