Real‐time fluorescence detection of multiple microscale cell culture analog devices in situ

Abstract

We investigated multiple microscale cell culture analog (lCCA) assays in situ with a high-throughput imaging system that provides quantitative, nondestructive, and realtime data on cell viability. Since samples do not move between measurements, captured images allow accurate time-course measurements of cell population response and tracking the fate of each cell type on a quantitative basis. The optical system was evaluated by measuring the short-term response to ethanol exposure and long-term growth of drug-resistant tumor cell lines with simultaneous samples. The optical system based on epi-fluorescent excitation consists of an LED and a CCD as well as discrete optical components for imaging a large number of cells simultaneously. HepG2/C3A and MESSA cell lines were cultured in two lCCA systems for continuous cell status monitoring in cell death experiments with ethanol and long-term cell growth. The experiment that tested ethanol uptake showed that ethanol immediately caused cell death. The system was applied to extracting dynamic constants in the uptake process. In the long-term cell growth experiment, growth of MESSA cells was followed by a stationary phase and eventual cell death attributed to nutrient and oxygen depletion and a change in the pH because of the accumulation of wastes by cell metabolism. HepG2/C3A cells were subject to contact inhibition and cell number did not change significantly over time. Issues related to long-term assays are also discussed. The quantitative results have been consistent with qualitative images and confirm the applicability of the portable optical system, and potential application to high-throughput analysis of cell-based assays to measure long-term dynamics. ' International Society for Analytical CytologyKey terms fluorescence detection; high-throughput screening; microscale cell culture analog; live cell imaging; cell status monitoring; green-fluorescent protein; cell dynamics A cell culture analog (CCA) device is a cell-based assay using interconnected compartments with different cell type and can be used to evaluate toxicity and efficacy of various pharmacological agents (1). Using the well-established semiconductor microfabrication technology, a microscale cell culture analog (lCCA) is fabricated from a silicon wafer. It consists of separate chambers for representing key organs in the human body, which are connected by microfluidic channels. The sizes of chambers and channels are designed so that they give similar residence times and liquid-to-cell ratios to the physiological values in the human body. A lCCA system has an advantage that it can mimic animal tissue dimensions more realistically than larger tissue culture vessels. This technology has been previously shown to replicate toxicological events that are undetectable using ordinary in vitro experiments (2,3). Microfluidic-based cell chips such as the lCCA presented in this paper are expected to provide a tool to decrease the time and cost of the drug discovery process especially in the phase prior to...

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