Ribonucleic acids were resolved by molecular sieve chromatography on columns of Sepharose 4B. The elution positions of messenger ribonucleic acids were determined by detection of polyadenosine tracts and by support of protein synthesis in a messenger-dependent cell-free system. The elution position of other ribonucleic acid species from the Sepharose 4B was determined by formamide-sucrose density gradient centrifugation. Resolution of ribonucleic acids by this column was not dependent on molecular weight but rather on other properties such as secondary structure or the presence of poly(adenylic acid). The elution profiles of ribonucleic acids on cross-linked Sepharose 4B differed markely from those on conventional Sepharose and appeared to depend on molecular size alone. There was diminished resolution of high molecular weight ribonucleic acids on such columns.