Methodology is described for the culture of avian and mammalian chondrocytes in ionotrophically gelled "semi-solid" and "hollow" alginate beads. Chondrocytes grown in "semi-solid" gels exhibited a spherical shape as opposed to a fibroblastic morphology observed in monolayer culture. In the "semi-solid" beads, the cells grew as small clumps and as large aggregates. The aggregates were round or elliptical in appearance and surrounded by a dense Alcian Blue positive halo. Preliminary studies with collagen and chitosan matrixes encapsulated in "hollow" beads suggest that cell growth and morphology are profoundly influenced by the composition of the cellular environment. Chondrocyte structure and function in the "semi-solid" and "hollow" beads were partially characterized by light microscopy, histochemical and biochemical means. The encapsulation methodology is readily applicable for the culture of chondrocytes in single beads, in multiwell dishes, or mass culture.
An improved method is described for the preparation of bovine testicular beta-galactosidase that allows the isolation of enzyme fractions that bind avidly to phosphomannosyl receptors. The procedure permits removal of a contaminating beta-hexosaminidase and yields nearly homogeneous beta-galactosidase. Enzyme eluted from DEAE-Sephacel was arbitrarily divided into pools that exhibited differing ability to bind phosphomannosyl receptors. A high binding fraction was rapidly assimilated by cultured cells and bound to both low and high molecular weight phosphomannosyl receptors. Carbohydrate analysis of the high binding fraction indicates an average content of one complex and one high mannose oligosaccharide chain per molecule and an average mannose 6-phosphate content of two residues per molecule. However, electrofocusing studies indicated that all the fractions were heterogeneous with respect to sialic acid and phosphate content. The purification procedure also provides highly purified beta-galactosidase suitable for removing beta-galactosidase residues from a variety of complex carbohydrates.
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