Rapid determination of general cell status, cell viability, and optimal harvest time in eukaryotic cell cultures by impedance flow cytometry

The determination of cell viability is essential to many areas of life sciences and biotechnology. Typically, cell viability measurements are based on the optical analysis of stained cells, which requires additional labeling steps and is hard to implement online. Frequency-dependent impedance flow cytometry (IFC) provides a label-free, fast, and reliable alternative to determine cell viability at the single cell level based on the Coulter principle. Here, we describe the application of IFC to eukaryotic cell cultures and compare the results to commonly used staining methods. Yeast cell parameters were assessed in normal and heat-inactivated cells as well as in alcoholic fermentation and long-term batch cultures providing a precise and fast determination of the cell viability and further quantitative measures of the cell culture status. As an important new application, we have investigated recombinant protein production in the widely used baculovirus insect cell expression system. The IFC analysis revealed the presence of a subpopulation of cells, which correlates with the protein expression yield, but it is not detectable with conventional optical cell counters. We tentatively identify this subpopulation as cells in the late phase of infection. Their detection can serve as a predictor for the optimal time point of harvest. The IFC technique should be generally applicable to many eukaryotic cell cultures in suspension, possibly also implemented online

Naturrasen: Planungsgrundlagen

Die neue Schrift bietet der Bauherrschaft, Planern und Betreibern, die erforderlichen Grundlagen zum erfolgreichen und nachhaltigen Bau und Betrieb von Naturrasen Spielfeldern

Tappeti erbosi naturali: Basi della pianificazione

Principi per una progettazione a regola d’arte: dimensioni di palestre, terreni da gioco e locali di servizio. Indicazioni su illuminazione, aerazione, acustica, etc

Gazons naturels: Bases de planification

La nouvelle publication fourni au maître d’ouvrage, planificateur et exploiteur les bases nécessaires pour la construction et l’exploitation de terrain de jeu en gazon naturel réussi et durable

Impedance Flow Cytometry: A Novel Technique in Pollen Analysis

<div><p>Introduction</p><p>An efficient and reliable method to estimate plant cell viability, especially of pollen, is important for plant breeding research and plant production processes. Pollen quality is determined by classical methods, like staining techniques or <i>in vitro</i> pollen germination, each having disadvantages with respect to reliability, analysis speed, and species dependency. Analysing single cells based on their dielectric properties by impedance flow cytometry (IFC) has developed into a common method for cellular characterisation in microbiology and medicine during the last decade. The aim of this study is to demonstrate the potential of IFC in plant cell analysis with the focus on pollen.</p><p>Method</p><p>Developing and mature pollen grains were analysed during their passage through a microfluidic chip to which radio frequencies of 0.5 to 12 MHz were applied. The acquired data provided information about the developmental stage, viability, and germination capacity. The biological relevance of the acquired IFC data was confirmed by classical staining methods, inactivation controls, as well as pollen germination assays.</p><p>Results</p><p>Different stages of developing pollen, dead, viable and germinating pollen populations could be detected and quantified by IFC. Pollen viability analysis by classical FDA staining showed a high correlation with IFC data. In parallel, pollen with active germination potential could be discriminated from the dead and the viable but non-germinating population.</p><p>Conclusion</p><p>The presented data demonstrate that IFC is an efficient, label-free, reliable and non-destructive technique to analyse pollen quality in a species-independent manner.</p></div