A soluble sialidase that can degrade recombinant glycoproteins expressed in Chinese hamster ovary (CHO) cells has been isolated and purified to near homogeneity from the cell culture fluid of this host. Purification of approximately 34,000-fold was carried out using conventional purification techniques including sequential DEAE-Sepharose and S-Sepharose ion-exchange chromatography, followed by hydrophobic interaction chromatography with Phenyl-Toyopearl. Final purification was achieved by heparin-agarose and chromatofocusing chromatography. The minimum molecular weight of the sialidase on SDS-PAGE was approximately 43,000 Da. When the final preparation was examined under non-denaturing conditions, two major (pI = 6.8 and 7.0) and five minor electrophoretic forms with different isoelectric points were identified. The basis for the electrophoretic heterogeneity is not known, but it was not due to carbohydrate diversity since no carbohydrates were detected on the purified protein. The enzyme degraded a variety of sialyl-conjugate substrates, at a pH optimum of 5.9, including intact glycoproteins, oligosaccharides and gangliosides with a 4-fold preference for 2,3- versus 2,6-linked sialic acid residues. With ganglioside substrates, internally linked sialic acid residues were not cleaved by the enzyme. Delineation of this enzyme from the lysosomal and plasma membrane sialidases was made using inhibition studies with C-9 substituted 5-acetamido-2,6-anhydro-3,5-dideoxy-D-glycero-D-galacto-non-2- enonic acid derivatives. The enzyme was identified in several CHO cell lines by immunoblotting using antiserum raised against a synthetic peptide based on amino acid sequence of a fragment derived by trypsin digestion of the purified sialidase.(ABSTRACT TRUNCATED AT 250 WORDS)

This paper presents a method of assigning tolerances to components so that (a) assemblies composed of the components meet specified functional requirements and (b) the cost of manufacturing all the components to their respective tolerance is minimised. Methods of obtaining tolerance equations from functional equations are briefly reviewed; the "method of extremes" is explained in detail. A relationship between cost and tolerance is derived, based upon published data.A method of assigning least-cost tolerances to a single tolerance equation is then developed.

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In this paper the effects of inspector error on a cost-based quality control system are investigated. The system examined is of a single sampling plan design involving several cost components. Both type I and type I1 inspector errors are considered. The model employs a process distribution, thus assuming that a stochastic process of some kind governs the quality of incoming lots. Optimal plan design is investigated under both error-free and error-prone inspection procedures and some comparisons are made.

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