Proteoglycan aggregates from bovine nasal cartilage, from Day 8 and Day 16 chondrocyte cultures derived from chick limb bud mesenchymal cells, and from em- bryonic chick epiphyseal cartilage were visualized as monolayers with electron microscopy. The lengths of the monomers in the aggregates and the average spac- ing of monomers on the central filament of hyaluronic acid were determined. The Gaussian distribution of core lengths for the bovine cartilage sample (343 + 95 nm) was significantly different from the distributions observed for the chick samples. With the exception of the sample derived from Day 16 cultures, all the chick samples gave statistically identical Gaussian distribu- tions (-310 f 60 nm). The Day 16 monomer core length distribution was somewhat skewed toward shorter lengths (297 + 58 nm). The average spacing of mono- mers on hyaluronic acid was larger for a reconstituted aggregate sample from Day 8 culture than for a prep- aration isolated from a similar culture by a procedure which did not involve dissociation of the aggregate structure. In all samples, the number of monomers per aggregate was linearly correlated with the length of hyaluronic acid in the central filament. Chondrocyte cultures were double-labeled with [3H]serine and [14C]serine on Day 8, or on Day 16, or both. Sepharose 2B analyses of the monomer fractions indicated that monomers synthesized on Day 16 had significantly smaller average sizes than those synthe- sized on Day 8. Further, although only 50% of the monomers synthesized on Day 8 remained in the tissue matrix on Day 16, the remaining molecules exhibited an identical size distribution to that of proteoglycans synthesized by Day 8 cultures and isolated directly. Differences in the average sizes of the chondroitin sul- fate chains in the monomer samples account in large part for the smaller sizes of the Day 16 macromolecules. Cartilage proteoglycan monomers consist of a protein core with covalently bound polysaccharide chains of chondroitin sulfate and keratan sulfate. The protein core of these macro- molecules can be divided into three regions; a chondroitin sulfate-enriched region, an intermediate keratan sulfate-en- riched region, and a hyaluronic acid binding region which is located at one end of the molecule (l-4). This noncovalent and highly specific interaction between the protein core of the * This work was supported in part (at Case Western Reserve University) by an Institutional Grant from the American Cancer Society, National Institutes of Health Grants HD-07209, 5T3-6M- 07225,HD-00020, HD-35609, AM-17110, and grants from the National Foundation (March of Dimes). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked