Optimising fed-batch production of recombinant proteins using the baculovirus expression vector system

Fed-batch culture can offer significant improvement in recombinant protein production compared to batch culture in the baculovirus expression vector system (BEVS), as shown by Nguyen et al. (1993) and Bedard et al. (1994) among others. However, a thorough analysis of fed-batch culture to determine its limits in improving recombinant protein production over batch culture has yet to be performed. In this work, this issue is addressed by the optimisation of single-addition fed-batch culture. This type of fed-batch culture involves the manual addition of a multi-component nutrient feed to batch culture before infection with the baculovirus. The nutrient feed consists of yeastolate ultrafiltrate, lipids, amino acids, vitamins, trace elements, and glucose, which were added to batch cultures of Spodoptera frugiperda (Sf9) cells before infection with a recombinant Autographa californica nuclear polyhedrosis virus (Ac-NPV) expressing beta-galactosidase (beta-Gal). The fed-batch production of beta-Gal was optimised using response surface methods (RSM). The optimisation was performed in two stages, starting with a screening procedure to determine the most important variables and ending with a central-composite experiment to obtain a response surface model of volumetric beta-Gal production. The predicted optimum volumetric yield of beta-Gal in fed-batch culture was 2.4-fold that of the best yields in batch culture. This result was confirmed by a statistical analysis of the best fed-batch and batch data (with average beta-Gal yields of 1.2 and 0.5 g/L, respectively) obtained from this laboratory. The response surface model generated can be used to design a more economical fed-batch operation, in which nutrient feed volumes are minimised while maintaining acceptable improvements in beta-Gal yield. (C) 1998 John Wiley & Sons, Inc.

The development of an early warning system to detect trends in illicit drug use in Australia: The illicit drug reporting-system

Appropriate ways to monitor the availability and use of illicit drugs were examined. Four methods were tested concurrently: (1) a quantitative survey of injecting drug users, (2) a qualitative key informant study of illicit drug users and professionals working in the drug field, (3) examination of existing sources of survey, health and law enforcement data and (4) an ethnographic study of a high risk group of illicit drug users. The first three methods were recommended for inclusion in an ongoing national monitoring system, enabling the collection of both quantitative and qualitative data on a range of illicit drugs in a relatively brief, quick and cost-effective manner. A degree of convergent validity was also noted among these methods, improving the degree of confidence in drug trends. The importance of injecting drug users as a sentinel population of illicit drug users was highlighted, along with optimal methods for qualitative research.

Expression of galectin-1 in the mouse olfactory system

Primary sensory olfactory axons arise from the olfactory neuroepithelium that lines the nasal cavity and then project via the olfactory nerve into the olfactory bulb. The P-galactoside binding lectin, galectin-1,and its laminin ligand have been implicated in the growth of these axons along this pathway. In galectin-1 null mutant mice, a subpopulation of primary sensory olfactory axons fails to reach its targets in the olfactory bulb. In the present study we examined the spatiotemporal expression pattern of galectin-1 in normal mice in order to understand its role in the development of the olfactory nerve pathway. At E15.5, when olfactory axons have already contacted the olfactory bulb, galectin-1 was expressed in the cartilage and mesenchyme surrounding the nasal cavity but was absent from the olfactory neuroepithelium, nerve and bulb. Between E16.5 and birth galectin-1 began to be expressed by olfactory nerve ensheathing cells in the lamina propria of the neuroepithelium and nerve fibre layer. Galectin-1 was neither expressed by primary sensory neurons in the olfactory neuroepithelium nor by their axons in the olfactory nerve. Laminin, a galectin-1 ligand, also exhibited a similar expression pattern in the embryonic olfactory nerve pathway. Our results reveal that galectin-1 is dynamically expressed by glial elements within the nerve fibre layer during a discrete period in the developing olfactory nerve pathway. Previous studies have reported galectin-1 acts as a substrate adhesion molecule by cross-linking primary sensory olfactory neurons to laminin. Thus, the coordinate expression of galectin-1 and laminin in the embryonic nerve fibre layer suggests that these molecules support the adhesion and fasciculation of axons en route to their glomerular targets.

A systemic secondary vessel system is present in the teleost fish Tandanus tandanus and absent in the elasmobranchs Carcharhinus melanopterus and Rhinobutos typus and in the dipnoan Neoceratodus forsteri

A system of secondary vessels emerging from the primary vessels as numerous coiled capillaries has been described in numerous teleost and holost fishes. The systemic secondary vessels of the teleost Tandanus tandanus are typical of this system and are described in this study. The existence of a secondary vessel system has been postulated in the elasmobranch group. No secondary vessel origins, as seen in the teleosts, are present in the elasmobranchs Rhinobatos typus and Carcharhinus melanopterus. Vessels with a similar distribution to secondary arteries are observed but these are venous rather than arterial in nature and do not connect with the primary arteries. Like the secondary veins in teleosts, the cutaneous veins in R. typus contain blood with a low haematocrit. There is no morphological evidence for a secondary vessel system in the dipnoan Neoceratodus forsteri.