The high-density lipoproteins (HDL) from canine, bovine, and chicken plasma have been shown to contain almost exclusively the apolipoprotein A-I, while human HDL contains a second major component, the apolipoprotein A-II. Chemical cross-linking demonstrated that dog and chicken HDL contain three apolipoprotein A-I molecules per particle, while bovine HDL contain approximately six apolipoprotein A-I molecules per particle. By this method, the amount of protein in human HDL2 (d = 1.063-1.12) was found to be approximately 120 000 g/mol, while for human HDL3 (d = 1.12-1.21) a value of approximately 90 000 g/mol was obtained, suggesting that the protein complement of HDL2 and HDL3 differ by only one apolipoprotein A-I chain per particle. Comparison of the apolipoprotein A-I from various animal species indicated that the canine and human apolipoprotein A-I proteins were the most similar by fluorescence, self-association properties, and immunoreactivity. Cross-linking of chicken and bovine apolipoprotein A-I yielded patterns distinctly different from that obtained with the human or canine counterpart. It is concluded that the quaternary structure of the various species of HDL is not directly correlated with the degree of self-association found for the protein constituents.