Dielectrophoretic analysis of the impact of isopropyl alcohol on the electric polarisability of Escherichia coli whole-cells.

Whole-cell biocatalysts are versatile tools in (industrial) production processes; though, the effects that impact the efficiency are not fully understood yet. One main factor that affects whole-cell biocatalysts is the surrounding medium, which often consists of organic solvents due to low solubility of substrates in aqueous solutions. It is expected that organic solvents change the biophysical and biochemical properties of the whole-cell biocatalysts, e.g. by permeabilising the cell membrane, and thus analysis of these effects is of high importance. In this work, we present an analysis method to study the impact of organic solvents on whole-cell biocatalysts by means of dielectrophoresis. For instance, we evaluate the changes of the characteristic dielectrophoretic trapping ratio induced by incubation of Escherichia coli, serving as a model system, in an aqueous medium containing isopropyl alcohol. Therefore, we could evaluate the impact on the electric polarisability of the cells. For this purpose, a special microchannel device was designed and Escherichia coli cells were genetically modified to reliably synthesise a green fluorescent protein. We could demonstrate that our method was capable of revealing different responses to small changes in isopropyl alcohol concentration and incubation duration. Complementary spectrophotometric UV-Vis (ultraviolet-visible light) absorbance analysis of released NAD(P)+/NAD(P)H cofactor and proteins confirmed our results. Based on our results, we discuss the biophysical effects taking place during incubation. Graphical abstract.

Reaching for the limits in continuous-flow dielectrophoretic DNA analysis.

The efficient purification and analysis of topological DNA variants is mandatory for many state-of-the-art molecular medicine technologies, like gene- and cancer-therapy as well as plasmid vaccination. In this work, we exploit dielectrophoresis (DEP) for a fast and efficient continuous-flow separation and analysis that goes beyond the standard methods of gel electrophoresis and capillary electrophoresis. The aim of this work was to reach for the limits in dielectrophoretic analysis of DNA regarding the size resolution and the topological conformation. A continuous-flow analytical separation of analyte mixtures of small linear DNA-fragments (10.0 kbp, 8.0 kbp, 6.0 kbp, and 5.0 kbp) and topological DNA variants (linear and supercoiled conformation) was investigated. We present a world record in the minimal size difference of 16.7% of DNA samples that can be resolved in a dielectrophoretic continuous-flow separation. Moreover, we demonstrate for the first time a microfluidic continuous-flow separation of DNA molecules based on their topological conformation. Since dielectrophoresis is virtually label-free, it offers a fast in-process quality control with low consumption, e.g. for the production of gene vaccines.