Continuously perfused, non-cross-contaminating microfluidic chamber array for studying cellular responses to orthogonal combinations of matrix and soluble signals

Abstract

We present a microfluidic cell culture array with unique versatility and parallelization for experimental trials requiring perfusion cultures. Specifically, we realize a rectangular chamber array in a PDMS device with three attributes: (i) continuous perfusion; (ii) flow paths that forbid cross-chamber contamination; and (iii) chamber shielding from direct perfusion to minimize shear-induced cell behaviour. These attributes are made possible by a bridge-and-underpass architecture, where flow streams travel vertically to pass over (or under) channels and on-chip valves. The array is also designed for considerable versatility, providing subarray, row, column, or single chamber addressing. It allows for incubation with adsorbed molecules, perfusion of differing media, seeding or extraction of cells, and assay staining. We use the device to characterize different phenotypes of alveolar epithelial type II (ATII) cells, particularly the extent of epithelial-to-mesenchymal transition (EMT), a highly suspected pathway in tissue regeneration and fibrosis. Cells are cultured on combinations of matrix proteins (fibronectin or laminin by row) and soluble signals (with or without transforming growth factor-β1 by column) with two repeats per chip. Fluorescent assays are performed in the array to assess viability, cytoskeletal organization, and cell–cell junction formation. Assay and morphological data are used to tease-out effects of cues driving each phenotype, confirming this as an effective and versatile combinatorial screening platform.