Abstract
Microfluidics, the science of engineering fluid streams at the micrometer scale, offers unique tools for creating and controlling gradients of soluble compounds. Gradient generation can be used to recreate complex physiological microenvironments, but is also useful for screening purposes. For example, in a single experiment, adherent cells can be exposed to a range of concentrations of the compound of interest, enabling high-content analysis of cell behaviour and enhancing throughput. In this study, we present the development of a microfluidic screening platform where, by means of diffusion, gradients of soluble compounds can be generated and sustained. This platform enables the culture of adherent cells under shear stress-free conditions, and their exposure to a soluble compound in a concentration gradient-wise manner. The platform consists of five serial cell culture chambers, all coupled to two lateral fluid supply channels that are used for gradient generation through a source-sink mechanism. Furthermore, an additional inlet and outlet are used for cell seeding inside the chambers. Finite element modeling was used for the optimization of the design of the platform and for validation of the dynamics of gradient generation. Then, as a proof-of-concept, human osteosarcoma MG-63 cells were cultured inside the platform and exposed to a gradient of Cytochalasin D, an actin polymerization inhibitor. This set-up allowed us to analyze cell morphological changes over time, including cell area and eccentricity measurements, as a function of Cytochalasin D concentration by using fluorescence image-based cytometry.
Highlights
In the past two decades, high-throughput screening (HTS) and high-content screening (HCS) have become major landmarks in the field of drug discovery, leading to fast identification of new therapeutic molecules and novel genetic engineering strategies (Zhao et al 2015; Lovitt et al 2013; CarlsonStevermer et al 2016; Macchi et al 2016)
A flow simulation was generated by Finite Element Modelling (FEM) for a cell culture chamber unit (Supplementary Fig. 1) to predict the concentration gradient profile of a soluble compound formed by diffusion between the Bsource^ and the Bsink^ channel
The results of this study showed that the platform developed here is suitable for tracking the changes in cell morphology over time, upon exposure to a concentration gradient created inside the microfluidic device over a monolayer of MG-63 cells
Summary
In the past two decades, high-throughput screening (HTS) and high-content screening (HCS) have become major landmarks in the field of drug discovery, leading to fast identification of new therapeutic molecules and novel genetic engineering strategies (Zhao et al 2015; Lovitt et al 2013; CarlsonStevermer et al 2016; Macchi et al 2016). By closely controlling fluid flows, microfluidic devices can be used to generate gradients of, for example, soluble molecules This capability can be exploited to expose cultured cells to a large range of concentrations of the compounds of interest in a single experiment (Harink et al 2015; Kilinc et al 2016; Xiao et al 2014; Zou et al 2015). The type of mechanism determines the profile of the gradient and its hydrodynamic characteristics inside the device (Berthier and Beebe 2014; Kim et al 2010)
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