Abstract A microsomal preparation from chick embryo epiphyseal cartilage has been used to study the assembly of the heteropolysaccharide, chondroitin. Endogenous chondroitin sulfate in the microsomal preparation was found to act as a primer for the addition of relatively short non-sulfated polysaccharide chains to form large molecules with a large sulfated portion and a small non-sulfated portion. A smaller primer was found to act as an acceptor for the polymerization of long non-sulfated chains to form large, predominantly non-sulfated chondroitin. Although the incorporation of sugars was time dependent, there was no detectable accumulation of short-chain molecules at the earliest times examined. The results suggest that the mechanism of polysaccharide formation is one of rapid assembly of most of the heteropolysaccharide chain, with subsequent addition taking place at a much slower rate. This would indicate a tightly organized mechanism of synthesis which may represent a general pattern for synthesis of heteropolysaccharides.