Evaluation of the impact of lipid fouling during the chromatographic purification of virus‐like particles from Saccharomyces cerevisiae

BACKGROUND: Production of recombinant virus‐like particles (VLPs) in yeast expression systems for use as vaccines requires cell disruption and detergent‐mediated steps to liberate the product. Typically, these release high levels of cellular components such as lipids that foul chromatography columns. This study compares the impact of applying lipid‐rich and lipid‐depleted feedstocks to hydrophobic interaction chromatography columns to quantify the loss of performance caused by the presence of host lipids over a total of 40 operational cycles. RESULTS: VLP binding capacity in the lipid‐rich feed was significantly lower than for the lipid‐depleted feed, with greater than 24% of the lipids remaining in the column after each cycle. Triacylglycerol was found to be the major contaminant. The effectiveness of subsequent caustic clean‐in‐place was limited, resulting in column hydrophobicity increasing over repeated loading cycles. This improved the effective VLP binding capacity and affinity, but also made product elution more difficult, and recovery decreased by more than 70% over the 40 cycles. CONCLUSION: Host cell lipids cause major fouling problems during VLP purification. Instead of screening for better CIP conditions, priority should be given to identifying suitable upstream lipid removal strategies in order to maintain column performance and so yield more economically viable processes. Copyright © 2009 Society of Chemical Industry     

Levels of functional impairment following a civilian disaster: The Beverly Hills Supper Club fire.

117 individuals (mean age 37.77 yrs) who were at the fire and 30 individuals (mean age 40.72 yrs) who were not at the fire (bereaved families and rescue workers) were assessed regarding objective stressfulness of their fire experience, subjective stressfulness, and intensity of psychological symptoms (the Psychiatric Evaluation Form, PEF) in a structured clinical interview approximately 1 yr after the fire. Ss also filled out the Symptom Checklist-90, Revised Version (SCL-90R). 88 Ss were followed-up at 2 yrs. The group as a whole was more impaired than comparison samples of normals but less impaired than outpatients. Ss at the fire were less impaired than those not at the fire, who were similar to outpatients on the PEF. The latter group improved significantly on several measures from 1 to 2 yrs postfire, whereas the group at the fire showed little change. Results are discussed in the context of the specific instruments and methodology used in the present study, impairment levels of other samples, and the nature of the particular disaster. (19 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Linguistic and location effects in compliance with pesticide warning labels for amateur and professional users

Three studies explored amateur and professional users' compliance with pesticide warning labels. Professionals were classified as people working in a profession in which the use of pesticides is a necessary part of their job. Amateurs used pesticides only in their leisure time. The first study showed that the wording used affected perception of the appropriateness of hazard statements, one of the most effective variations being the use of the personal pronoun (statements beginning "You should..."). The location of warning information was also found to affect actual compliance: Compliance increased when warning information was presented in the directions for use section. A supplemental directive increased compliance only for professional users. In a final study, "best-case" and "worst-case" linguistic variations were combined with best-case and worst-case locations for safety information. Instruction statements using the personal pronoun and presented in the directions for use section resulted in the highest levels of compliance. The differences in compliance between amateur and professional users are interpreted within the framework of Rasmussen's (1986) distinction among skill-, rule-, and knowledge-based behavior. Actual or potential applications of this research include the design of warning labels and safety information.     

Power spectral densities of bit and word-interleaved line-coded channels for Gbit/s transmission

General formulas are presented for the power spectral densities (PSDs) of bit and word-interleaved line-coded digital signals, which are useful in the design of Gbit/s transmission systems. It is shown that word interleaving introduces very little spectral degradation for commonly used line codes, whereas the effects of bit-interleaving, though marked, are simply quantified. These predictions are confirmed by calculations of the PSDs using Monte Carlo simulations.<>     

Heterogeneity in auditory alarm sets makes them easier to learn

The primary objective of the experiments reported here was to demonstrate the effects of opening up the design envelope for auditory alarms on the ability of people to learn the meanings of a set of alarms. Two sets of alarms were tested, one already extant and one newly-designed set for the same set of functions, designed according to a rationale set out by the authors aimed at increasing the heterogeneity of the alarm set and incorporating some well-established principles of alarm design. For both sets of alarms, a similarity-rating experiment was followed by a learning experiment. The results showed that the newly-designed set was judged to be more internally dissimilar, and easier to learn, than the extant set. The design rationale outlined in the paper is useful for design purposes in a variety of practical domains and shows how alarm designers, even at a relatively late stage in the design process, can improve the efficacy of an alarm set.     

Visualising bioprocesses using ‘3D‐Windows of Operation’

The operation of bioprocesses involves handling many variables with complex interactions. The visualisation of interactions is vital if suitable conclusions are to be drawn. This paper extends the previously developed ‘Windows of Operation’ to three dimensions, enabling the effects of three parameters to be visualised simultaneously. A tool with a graphical user interface has been developed to enable the generation of ‘3D‐Windows of Operation’ and the visualisation of the effects of the variation of any three control variables. Its use is illustrated with simulations of enzyme production. A number of control variables and the effects of the imposition of constraints on the range of possible operating values are investigated. Changes in the range possible are related to the variation in specific constraints or variables. Finally, it is illustrated how these diagrams and operation heuristics may be used to select suitable operating conditions for the bioprocess. Copyright © 2004 Society of Chemical Industry     

The synchronization of variable-length codes

Many variable-length codes exhibit a tendency for resynchronization to occur automatically following any error. However, attempts to identify an underlying synchronization mechanism, and to accurately predict the expected synchronization delay, for even quite specific variable-length codes, appear to have been largely unsuccessful. The present paper explores a novel method for estimating the synchronization performance for a wide variety of variable-length codes, based on the T-Codes. T-Codes are a class of self-synchronizing codes, which typically synchronize within 2-3 codewords by a mechanism that derives from a recursive T-augmentation construction. It is observed that the T-Code mechanism for synchronization is followed, more or less, by other variable-length codes wherever substantial numbers of codewords are shared with a T-Code set. T-augmentation itself provides a means for assessing the contribution individual codewords make to the overall synchronization process for a T-Code set. Thus codeword differences between sets may be specifically evaluated to estimate the synchronization performance of a variable-length code set from a closely related T-Code set     

Some aspects of the static and dynamic compressive behaviour of β-brass single crystals

The compressive behaviour of β-brass single crystals has been investigated in uninterrupted static and dynamic tests and in interrupted tests using static-static, static-dynamic, dynamic-dynamic and dynamic-static loading sequences. Static and dynamic strain-rates were 1.5×10^?4 and 3.2×10³ sec^?1 respectively. Slip traces on the statically deformed crystals were wavy and deformation occurred by single slip on either (ī01) [111] or (¯211) [111] or by a transition mode involving both (ī01) [111] and (¯211 [111]. Except for anomalous behaviour in the dynamic reload following static preload the dynamic slip traces were straight with deformation occurring by multiple slip on four {110} planes involving two 〈111〉 directions. It is shown that there is no direct causal relationship between the lower work-hardening rate and level of flow stress and the crystallography of slip in dynamic deformation. The work-hardening rate and flow stress in static and dynamic loading are rather determined by the dynamics of the deformation. The differences in the substructural features as observed by transmission electron microscopy arise principally from the differences in the slip modes and cannot be interpreted as controlling the stress-strain behaviour. The low work-hardening rate and flow stress in dynamic deformation is believed to be due to the production of short-lived disorder. The absence of a/2〈111〉 dislocations in thin foils is explained in terms of the fast reordering reaction in β-brass.     

Study of the conditions for multi‐modal chromatographic capture of Fab′ from dual‐salt precipitated E. coli homogenate

BACKGROUND: In the commercial‐scale purification of immunotherapeutics, Protein A chromatography is employed routinely for its high binding capacity and selectivity. Nevertheless, matrix cost and ligand leaching issues remain and there are many alternatives such as ion‐exchange and multi‐modal resins that are less expensive. However, binding capacities are lower than Protein A owing to the co‐adsorption of protein impurities. One solution involves removing impurities before chromatography by precipitation and a potential approach presented in the literature recently employs a dual‐salt precipitation technique. The current study explores the impact of this upon the capture of an antibody fragment by a multimodal cation exchange resin. RESULTS: The dual salt precipitation procedure employed here removed 36% of the contaminant proteins. A microscale chromatography pipette tip approach was used in a high throughput screening format to scout rapidly for favourable binding conditions successfully. Higher binding capacities were achieved by activating the hydrophobic binding modality of the resin at feed ionic strengths of around 150 mS cm^?1 using ammonium sulphate. This was better than using the ion‐exchange modality at below 10 mS cm^?1. CONCLUSIONS: Dual salt precipitation followed by chromatographic capture using this resin were found to be complementary to one another since the removal of protein impurities by dual salt precipitation resulted in a two‐fold improvement in terms of binding capacity. © 2012 Society of Chemical Industry