Relation between cell disruption conditions, cell debris particle size, and inclusion body release

The efficiency of physical separation of inclusion bodies from cell debris is related to cell debris size and inclusion body release and both factors should be taken into account when designing a process. In this work, cell disruption by enzymatic treatment with lysozyme and cellulase, by homogenization, and by homogenization with ammonia pretreatment is discussed. These disruption methods are compared on the basis of inclusion body release, operating costs, and cell debris particle size. The latter was measured with cumulative sedimentation analysis in combination with membrane-associated protein quantification by SDS-PAGE and a spectrophotometric pepticloglycan quantification method. Comparison of the results obtained with these two cell debris quantification methods shows that enzymatic treatment yields cell debris particles with varying chemical composition, while this is not the case with the other disruption methods that were investigated. Furthermore, the experiments show that ammonia pretreatment with homogenization increases inclusion body release compared to homogenization without pretreatment and that this pretreatment may be used to control the cell debris size to some extent. The enzymatic disruption process gives a higher product release than homogenization with or without ammonia pretreatment at lower operating costs, but it also yields a much smaller cell debris size than the other disruption process. This is unfavorable for centrifugal inclusion body purification in this case, where cell debris is the component going to the sediment and the inclusion body is the floating component. Nevertheless, calculations show that centrifugal separation of inclusion bodies from the enzymatically treated cells gives a high inclusion body yield and purity. (C) 2004 Wiley Periodicals, Inc.

Particle bed charge decay behaviour under high tension roll separation

A new theory of particle discharge in high tension roll (HTR) separation is presented. The discharge dynamics of an isolated charged particle resting on a conducting surface are studied first. The analysis is extended to particle discharge in a homogenous particle bed. Finally, the paper looks at the more realistic scenario of particle discharge in a non-homogenous particle bed. The consequences of the resulting theory on HTR separation are discussed. Predictions from the new theory are tested against experimental HTR separations at the pilot scale. (c) 2005 Elsevier Ltd. All rights reserved.

Engineering of needle-free physical methods to target epidermal cells for DNA vaccination

A challenge in epidermal DNA vaccination is the efficient and targeted delivery of polynucleotides to immunologically sensitive Langerhans cells. This paper investigates this particular challenge for physical delivery approaches. The skin immunology and material properties are examined in the context of the physical cell targeting requirements of the viable epidermis. Selected current physical cell targeting technologies engineered to meet these needs are examined: needle and syringe; diffusion patches; liquid jet injectors; microneedle arrays/patches; and biolistic particle injection. The operating methods and relative performance of these approaches are discussed, with a comment on potential future developments and technologies. (c) 2005 Elsevier Ltd. All rights reserved.

Particle tracking in a salinity gradient: A method for measuring sinking rate of individual phytoplankton in the laboratory

This paper presents a new method to measure the sinking rates of individual phytoplankton “particles” (cells, chains, colonies, and aggregates) in the laboratory. Conventional particle tracking and high resolution video imaging were used to measure particle sinking rates and particle size. The stabilizing force of a very mild linear salinity gradient (1 ppt over 15 cm) prevented the formation of convection currents in the laboratory settling chamber. Whereas bulk settling methods such as SETCOL provide a single value of sinking rate for a population, this method allows the measurement of sinking rate and particle size for a large number of individual particles or phytoplankton within a population. The method has applications where sinking rates vary within a population, or where sinking rate-size relationships are important. Preliminary data from experiments with both laboratory and field samples of marine phytoplankton are presented here to illustrate the use of the technique, its applications, and limitations. Whereas this paper deals only with sinking phytoplankton, the method is equally valid for positively buoyant species, as well as nonbiological particles.