Droplet-based cell viability assay for analysis of spheroid formation, proliferation and high-resolution IC50 profiling

Abstract

Three-dimensional (3D) cell cultures or samples generated from biopsies are typically used as patient-specific in vitro models. As 3D cell cultures form cell–cell and cell–matrix interactions and mimic the in vivo situation better compared to monolayer cultures, they provide more reliable data for drug screening applications. In the field of drug screening, microfluidics is moving to the forefront for testing the efficacy of drugs, as measured by IC₅₀ values. Droplet-based microfluidics not only shares the advantages of well plate-based systems but also those that go beyond. The high-throughput character of droplet-based microfluidics enables the generation of hundreds of droplets per minute, with smaller volumes than in well plate-based systems. The high level of automation and the closed character of such systems permit a higher reproducibility of the generated data, as the well-known problem of evaporation in well plates is negligible. In this study, a modular droplet-based microfluidic platform is introduced that facilitates the formation of 3D cell cultures. For assessing cell viability in spheroids of the human embryonic kidney cell line, HEK-293, a resazurin-based CellTiter-Blue® assay was established on a droplet-based platform. Here, the pipe based bioreactors (pbb) technology was used to create a continuous drug gradient, enabling the realisation of 290 concentration levels within a single droplet sequence to determine high-resolution IC₅₀ values. Consequently, the pbb technology exceeds the state of the art, as only discrete concentrations of drugs are investigated in well plate-based systems. DMSO was used for drug testing experiments, as drugs are typically dissolved in it. As it is important that healthy cells are not affected by the drug or its solvents, the influence of DMSO was examined. Overall, the presented platform not only offers a robust and precise tool for validating drug efficacy using 3D cell cultures but also provides the basis for developing innovative therapies across a wide range of diseases. The modular design of the pbb platform provides the flexibility to address a variety of biomedical applications, ultimately accelerating personalized medicine to deliver better outcomes for patients.