Phenotypic Pattern-Based Assay for Dynamically Monitoring Host Cellular Responses to Salmonella Infections

Abstract

The interaction between mammalian host cells and bacteria is a dynamic process, and the underlying pathologic mechanisms are poorly characterized. Limited information describing the host-bacterial interaction is based mainly on studies using label-based endpoint assays that detect changes in cell behavior at a given time point, yielding incomplete information. In this paper, a novel, label-free, real-time cell-detection system based on electronic impedance sensor technology was adapted to dynamically monitor the entire process of intestinal epithelial cells response to Salmonella infection. Changes in cell morphology and attachment were quantitatively and continuously recorded following infection. The resulting impedance-based time-dependent cell response profiles (TCRPs) were compared to standard assays and showed good correlation and sensitivity. Biochemical assays further suggested that TCRPs were correlated with cytoskeleton-associated morphological dynamics, which can be largely attenuated by inhibitions of actin and microtubule polymerization. Collectively, our data indicate that cell-electrode impedance measurements not only provide a novel, real-time, label-free method for investigating bacterial infection but also help advance our understanding of host responses in a more physiological and continuous manner that is beyond the scope of current endpoint assays.

Association between phospholipase C epsilon gene (PLCE1) polymorphism and colorectal cancer risk in a Chinese population

“…Three replicates were performed for each sample and the quantitative real-time PCR products were analysed using Bio-Rad CFX Manager TM software, version 3.0 (Bio-Rad) using the 2 ÀÁÁCT method. 31 …”

Effects of Calcium on the Interactions of Acinetobacter baumannii with Human Respiratory Epithelial Cells

“…Compared to traditional endpoint detection, RTCA has the advantages of being non-invasive and highly accurate, as well as providing real-time monitoring, complete TCRPs, and easy operation. It is widely used in cytology research, such as in cell migration and invasion assays, cytotoxicity tests, gene regulation and cell-microenvironment interactions[15,[19][20][21]. Therefore, the obtained TCRPs can provide better information on the effects of calcium on host-bacterial interactions, complementing the results of our microscopy observations.…”

“…A label-free and noninvasive detection system (RTCA S16 system, ACEA Biosciences Inc.) based on dynamical and quantitative monitoring of cellular impedance in real time can produce specific timedependent cell response profile (TCRPs) patterns. This approach can provide biological information related to cellular physiological function for studying host-bacterial interactions [15]. Thus, in this work, we developed microscopic imaging, quantitative PCR (qPCR) and TCRP methods for continuously monitoring the interactions between Ab and human respiratory epithelial cells and the effects of calcium on host-bacterial interactions.…”

A Real-Time Impedance Based Method to Assess Rhodococcus equi Virulence

“…Although efficient mutagenesis tools for R. equi have been developed [12]–[14], the analysis of the resulting mutants with regard to their virulence is time consuming. The aim of this work was to demonstrate that measurement of impedance changes in cell culture wells using the xCELLigence system [22], [23], [28] can be used as a high-throughput system to analyse virulence of R. equi in real-time and that any effects recorded could be correlated to visual changes in the host cells.…”

“…A major advantage of this system is that it allows label-free analysis of the effects of experimental manipulation on the cell response in real-time [18]–[21]. Recently, an impedance based system was used to analyse the response of host cells to infection by the human pathogens Neisseria meningitidis and Salmonella typhimurium [22], [23].…”