Reproducibility and robustness of a real-time microfluidic cell toxicity assay

Numerous opportunities exist to apply microfluidic technology to high-throughput and high-content cell-based assays. However, maximizing the value of microfluidic assays for applications such as drug discovery, screening, or toxicity evaluation will require assurance of within-device repeatability, day-to-day reproducibility, and robustness to variations in conditions that might occur from laboratory to laboratory. This report describes a study of the performance and variability of a cell-based toxicity assay in microfluidic devices made of poly(dimethylsiloxane) (PDMS). The assay involves expression of destabilized green fluorescent protein (GFP) as a reporter of intracellular protein synthesis and degradation. Reduction in cellular GFP due to inhibition of ribosome activity by cycloheximide (CHX) was quantified with real-time quantitative fluorescence imaging. Assay repeatability was measured within a 64-chamber microfluidic device. Assay performance across a range of cell loading densities within a single device was assessed, as was replication of measurements in microfluidic devices prepared on different days. Assay robustness was tested using different fluorescence illumination sources and reservoir-to-device tubing choices. Both microfluidic and larger scale assay conditions showed comparable GFP decay rates upon CHX exposure, but the microfluidic data provided the higher level of confidence.