Proteins of cytoplasmic ribosomes from human tonsillar lymphatic tissue were analyzed by twodimensional polyacrylamide-gel electrophoresis. 34 proteins were found to be associated with the 404 subunit and 35 proteins with the 60-S subunit. 65 proteins had different electrophoretic mobilities. Two proteins found in the small subunit (SIO and S20) co-migrated with two large subunit proteins (L13 and L20 respectively). A comparison of our results with those obtained by the same method for ribosomal proteins from human placenta, rat liver and rabbit reticulocytes suggested a possible identity of the electrophoretic mobilities of 21 -24 40-S subunit proteins and 25-26 6 0 4 subunit proteins.Proteins of cytoplasmic ribosomes from various eucaryotic species and within the same species from various tissues have recently been analyzed by twodimensional gel electrophoresis [l-131. These studies revealed a considerable similarity between the eucaryotic ribosomal proteins of different species. However, some interspecific and also intraspecific differences have been observed [5 -7,111. This communication will report the separation of ribosomal proteins from human tonsillar lymphatic tissue using the method of Kaltschmidt and Wittman MATERIALS AND METHODS ChemicalsAcrylamide, methylene-N,W-bisacrylamide were obtained from Biomol (Ilvesheim) ; urea, trizma base from Baker (Deventer); N,N,N',N'-tetramethylethylendiamine from Serva (Heidelberg). All the other reagent-grade chemicals were purchased from Merck (Darmstadt). Preparations of Ribosomes and Extraction of Ribosomal ProteinsRibosomes [15] and ribosomal subunits [16] from human tonsillar lymphatic tissue were prepared as previously described. Ribosomal proteins were extracted according to the method of Spitnik-Elson [17]. 80-S ribosomes corresponding to 100 ,4260 units, 60-S ribosomal subunits corresponding to 60 A,,, units or 40-S ribosomal subunits corresponding to 40 ,4260 units were suspended in 75 pl of medium A minus sucrose (50 mM Tris-HC1 pH 7.4,5 mM MgC12, 25 mM KCl, and 7 mM 2-mercaptoethanol) in 2-ml tubes and mixed with an equal volume of 4 M LiCl in 8 M urea. After 24 h at 4 "C the RNA precipitate was pelleted by centrifugation for 10 min at 27000 x g in a Sorvall SS-34 rotor. The pellet was resuspended in 50 pl of medium A minus sucrose and extracted again. The supernatant fractions were combined and dialyzed for 30 h against several changes of sample gel solution ~4 1 . Two-Dimensional Electrophoresis and Numbering of Ribosomal ProteinsTwo-dimensional polyacrylamide-gel electrophoresis was performed according to the method of KaltEur. J. Biochem. 50 (1974)