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Roche’s xCELLigence MP Instrument Used in the First Human Kinome Screen for Dynamic RNA Inhibitor Phenotypes During Cell Proliferation

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RNA interference studies are a powerful technology for high-throughput screening. However, endpoint analyses only provide a snapshot of the analyzed cellular phenotype, as it is not possible to measure developmental changes over time.

To address this, a research team led by Stefan Wiemann and Ulrich Tschulena in the Division Molecular Genome Analysis of the German Cancer Research Center (DKFZ) in Heidelberg, Germany used the xCELLigence MP Instrument, from Roche, to quantify cell proliferation, without the need for tagging or modifying sampled cells.

As reported in the July 2011 (Zhang et al., Vol. 6, Issue 7) of PLoS ONE Online, they showed that the measurement of impedance strength is positively correlated with the number of cells attached to electrodes.

Importantly, impedance measurements reflect not only cell number and also the quality of the cells’ interaction with their substrate, making this technology a sensitive and reliable way to assess cell status, including cell morphology, cell adhesion and cell viability.

The Heidelberg German Cancer Center research team integrated the Roche xCELLigence MP Instrument into a high-throughput workflow for assaying the effects of large numbers of different small interfering RNA (siRNA) transfections.

They used a human siRNA library to perform a whole kinome screen, targeting 779 kinases and 80 cell cycle genes to analyze cell proliferation in real time, by monitoring the dynamics of the cellular responses after high-throughput gene knockdown.

The team’s findings showed that xCELLigence System measurements correlate with results obtained using, in parallel, conventional endpoint analyses, in particular Promega’s CellTiter-Blue and Roche’s WST-1 Cell Proliferation Reagent for assaying cell viability, as well as qRT-PCR for quantifying gene expression.

However, the dynamic data obtained with the xCELLigence System allowed to identify the timing after transfection where the effect was maximal and to sort genes according to these time points.

“We have carried out a human kinome RNAi screen using xCELLigence with electrical impedance as output. This screen has confirmed previously identified inhibitor genes, as well as activators of cell proliferation. Our data establish the technology of the xCELLigence system as a novel tool amenable for high-throughput screening, opening new avenues in the dynamic cellular analysis of phenotypes induced by RNAi and other perturbations,” summarizes Stefan Wiemann, one of the authors of the study and Head of the Division Molecular Genome Analysis at DKFZ.

The xCELLigence MP Instrument uses proprietary software and E-Plates 96 to measure electronic cell impedance using sensor electrodes. Computer-controlled signal generation and automatic frequency scanning, enable continual precise detection of changes in cell behavior using 96-well culture plates.