The application of pulsed electric fields (PEFs) is becoming a promising tool for application in biotechnology, and the food industry. However, real-time monitoring of the efficiency of PEF treatment conditions is challenging, especially at the industrial scale and in continuous production conditions. To overcome this challenge, we have developed a straightforward setup capable of real-time detection of yeast biological autoluminescence (BAL) during pulsing. Saccharomyces cerevisiae culture was exposed to 8 pulses of 100 µs width with electric field strength magnitude 2-7 kV cm-1. To assess the sensitivity of our method in detecting yeast electroporation, we conducted a comparison with established methods including impedance measurements, propidium iodide uptake, cell growth assay, and fluorescence microscopy. Our results demonstrate that yeast electroporation can be instantaneously monitored during pulsing, making it highly suitable for industrial applications. Furthermore, the simplicity of our setup facilitates its integration into continuous liquid flow systems. Additionally, we have established quantitative indicators based on a thorough statistical analysis of the data that can be implemented through a dedicated machine interface, providing efficiency indicators for analysis.
- Klíčová slova
- autoluminescence, electroporation, pulsed electric fields, yeast in biotechnology,
- MeSH
- elektroporace * metody MeSH
- Saccharomyces cerevisiae * růst a vývoj MeSH
- Publikační typ
- časopisecké články MeSH
Pulsed electric field (PEF) technology is promising for the manipulation of biomolecular components and has potential applications in biomedicine and bionanotechnology. Microtubules, nanoscopic tubular structures self-assembled from protein tubulin, serve as important components in basic cellular processes as well as in engineered biomolecular nanosystems. Recent studies in cell-based models have demonstrated that PEF affects the cytoskeleton, including microtubules. However, the direct effects of PEF on microtubules are not clear. In this work, we developed a lab-on-a-chip platform integrated with a total internal reflection fluorescence microscope system to elucidate the PEF effects on a microtubules network mimicking the cell-like density of microtubules. The designed platform enables the delivery of short (microsecond-scale), high-field-strength ([Formula: see text] 25 kV/cm) electric pulses far from the electrode/electrolyte interface. We showed that microsecond PEF is capable of overcoming the non-covalent microtubule bonding force to the substrate and translocating the microtubules. This microsecond PEF effect combined with macromolecular crowding led to aggregation of microtubules. Our results expand the toolbox of bioelectronics technologies and electromagnetic tools for the manipulation of biomolecular nanoscopic systems and contribute to the understanding of microsecond PEF effects on a microtubule cytoskeleton.
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Normal or excessive oxidative metabolism in organisms is essential in physiological and pathophysiological processes, respectively. Therefore, monitoring of biological oxidative processes induced by the chemical or physical stimuli is nowadays of extreme importance due to the environment overloaded with various physicochemical factors. Current techniques typically require the addition of chemical labels or light illumination, which perturb the samples to be analyzed. Moreover, the current techniques are very demanding in terms of sample preparation and equipment. To alleviate these limitations, we propose a label-free monitoring tool of oxidation based on biological autoluminescence (BAL). We demonstrate this tool on Saccharomyces cerevisiae cell culture. We showed that BAL can be used to monitor chemical perturbation of yeast due to Fenton reagents initiated oxidation-the BAL intensity changes with hydrogen peroxide concentration in a dose-dependent manner. Furthermore, we also showed that BAL reflects the effects of low-frequency magnetic field on the yeast cell culture, where we observed a disturbance of the BAL kinetics in the exposed vs. control case. Our results contribute to the development of novel techniques for label-free, real-time, noninvasive monitoring of oxidative processes and approaches for their modulation.
- MeSH
- celulosa analogy a deriváty farmakologie MeSH
- fixní kombinace léků MeSH
- kultivační techniky MeSH
- luminiscence * MeSH
- oxidace-redukce účinky léků MeSH
- povidon farmakologie MeSH
- Saccharomyces cerevisiae cytologie účinky léků metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- celulosa MeSH
- fixní kombinace léků MeSH
- hyetellose, povidone drug combination MeSH Prohlížeč
- povidon MeSH
